Purpose: Cancer stem cells (CSC) are thought to represent the population of tumorigenic cells responsible for tumor development. The stem cell antigen CD133 identifies such a tumorigenic population in a subset of glioblastoma patients. We conducted a prospective study to explore the prognostic potential of CSC analysis in glioblastoma patients. Experimental Design: We investigated the relationship between the in vitro growth potential of glioblastoma CSCs and patient death or disease progression in tumors of 44 consecutive glioblastoma patients treated with complete or partial tumorectomy followed by radiotherapy combined with temozolomide treatment. Moreover, we evaluated by immunohistochemistry and immunofluorescence the prognostic value of the relative presence of CD133⁺ and CD133⁺/Ki67⁺ cells in patient tumors. Results:In vitro CSC generation and the presence of ≥2% CD133⁺ cells in tumor lesions negatively correlated with overall (P = 0.0001 and 0.02, respectively) and progression-free (P = 0.0002 and 0.01, respectively) survival of patients. A very poor overall (P = 0.007) and progression-free (P = 0.001) survival was observed among patients whose tumors contained CD133⁺ cells expressing Ki67. Taking into account symptom duration, surgery type, age, O⁶-methylguanine-DNA methyltransferase promoter methylation, and p53 status, generation of CSCs and CD133/Ki67 coexpression emerged as highly significant independent prognostic factors, with an adjusted hazard ratio of 2.92 (95% confidence interval, 1.37-6.2; P = 0.005) and 4.48 (95% confidence interval, 1.68-11.9; P = 0.003), respectively. Conclusions: The analysis of CSCs may predict the survival of glioblastoma patients. In vitro CSC generation and presence of CD133⁺/Ki67⁺ cells are two considerable prognostic factors of disease progression and poor clinical outcome.

Mesenchymal stromal cells may represent an ideal candidate to deliver anti-cancer drugs. In a previous study, we demonstrated that exposure of mouse bone marrow derived stromal cells to Doxorubicin led them to acquire anti-proliferative potential towards co-cultured haematopoietic stem cells (HSCs). We thus hypothesized whether freshly isolated human bone marrow Mesenchymal stem cells (hMSCs) and mature murine stromal cells (SR4987 line) primed in vitro with anti-cancer drugs and then localized near cancer cells, could inhibit proliferation. Paclitaxel (PTX) was used to prime culture of hMSCs and SR4987. Incorporation of PTX into hMSCs was studied by using FICT-labelled-PTX and analyzed by FACS and confocal microscopy. Release of PTX in culture medium by PTX primed hMSCs (hMSCsPTX) was investigated by HPLC. Culture of Endothelial cells (ECs) and aorta ring assay were used to test the anti-angiogenic activity of hMSCsPTX and PTX primed SR4987(SR4987PTX), while anti-tumor activity was tested in vitro on the proliferation of different tumor cell lines and in vivo by co-transplanting hMSCsPTX and SR4987PTX with cancer cells in mice. Nevertheless, despite a loss of cells due to chemo-induced apoptosis, both hMSCs and SR4987 were able to rapidly incorporate PTX and could slowly release PTX in the culture medium in a time dependent manner. PTX primed cells acquired a potent anti-tumor and anti-angiogenic activity in vitro that was dose dependent, and demonstrable by using their conditioned medium or by co-culture assay. Finally, hMSCsPTX and SR4987PTX co-injected with human cancer cells (DU145 and U87MG) and mouse melanoma cells (B16) in immunodeficient and in syngenic mice significantly delayed tumor takes and reduced tumor growth. These data demonstrate, for the first time, that without any genetic manipulation, mesenchymal stromal cells can uptake and subsequently slowly release PTX. This may lead to potential new tools to increase efficacy of cancer therapy.

Glioma stem‐like cells (GSCs) correspond to a tumor cell subpopulation, involved in glioblastoma multiforme (GBM) tumor initiation and acquired chemoresistance. Currently, drug‐induced differentiation is considered as a promising approach to eradicate this tumor‐driving cell population. Recently, the effect of cannabinoids (CBs) in promoting glial differentiation and inhibiting gliomagenesis has been evidenced. Herein, we demonstrated that cannabidiol (CBD) by activating transient receptor potential vanilloid‐2 (TRPV2) triggers GSCs differentiation activating the autophagic process and inhibits GSCs proliferation and clonogenic capability. Above all, CBD and carmustine (BCNU) in combination overcome the high resistance of GSCs to BCNU treatment, by inducing apoptotic cell death. Acute myeloid leukemia (Aml‐1) transcription factors play a pivotal role in GBM proliferation and differentiation and it is known that Aml‐1 control the expression of several nociceptive receptors. So, we evaluated the expression levels of Aml‐1 spliced variants (Aml‐1a, b and c) in GSCs and during their differentiation. We found that Aml‐1a is upregulated during GSCs differentiation, and its downregulation restores a stem cell phenotype in differentiated GSCs. Since it was demonstrated that CBD induces also TRPV2 expression and that TRPV2 is involved in GSCs differentiation, we evaluated if Aml‐1a interacted directly with TRPV2 promoters. Herein, we found that Aml‐1a binds TRPV2 promoters and that Aml‐1a expression is upregulated by CBD treatment, in a TRPV2 and PI3K/AKT dependent manner. Altogether, these results support a novel mechanism by which CBD inducing TRPV2‐dependent autophagic process stimulates Aml‐1a‐dependent GSCs differentiation, abrogating the BCNU chemoresistance in GSCs.