Cancer stem cells (CSCs) or cancer initiating cells (CICs) maintain self-renewal and multilineage differentiation properties of various tumors, as well as the cellular heterogeneity consisting of several subpopulations within tumors. CSCs display the malignant phenotype, self-renewal ability, altered genomic stability, specific epigenetic signature, and most of the time can be phenotyped by cell surface markers (e.g., CD133, CD24, and CD44). Numerous studies support the concept that non-stem cancer cells (non-CSCs) are sensitive to cancer therapy while CSCs are relatively resistant to treatment. In glioblastoma stem cells (GSCs), there is clonal heterogeneity at the genetic level with distinct tumorigenic potential, and defined GSC marker expression resulting from clonal evolution which is likely to influence disease progression and response to treatment. Another level of complexity in glioblastoma multiforme (GBM) tumors is the dynamic equilibrium between GSCs and differentiated non-GSCs, and the potential for non-GSCs to revert (dedifferentiate) to GSCs due to epigenetic alteration which confers phenotypic plasticity to the tumor cell population. Moreover, exposure of the differentiated GBM cells to therapeutic doses of temozolomide (TMZ) or ionizing radiation (IR) increases the GSC pool both in vitro and in vivo. This review describes various subtypes of GBM, discusses the evolution of CSC models and epigenetic plasticity, as well as interconversion between GSCs and differentiated non-GSCs, and offers strategies to potentially eliminate GSCs.

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Fanconi anemia is a genetic disorder characterized by bone marrow failure. Significant evidence supports enhanced apoptosis of hematopoietic stem/progenitor cells as a critical factor in the pathogenesis of bone marrow failure in Fanconi anemia. However, the molecular mechanism(s) responsible for the apoptotic phenotype are incompletely understood. Here, we tested whether alterations in the activation of a redox-dependent pathway may participate in the pro-apoptotic phenotype of primary Fancc ؊/؊ cells in response to oxidative stress. Our data indicate that Fancc ؊/؊ cells are highly sensitive to oxidant stimuli and undergo enhanced oxidant-mediated apoptosis compared with wild type controls. In addition, antioxidants preferentially enhanced the survival of Fancc ؊/؊ cells. Because oxidative stress activates the redox-dependent ASK1 pathway, we assessed whether Fancc ؊/؊ cells exhibited increased oxidant-induced ASK1 activation. Our results revealed ASK1 hyperactivation in H 2 O 2 -treated Fancc ؊/؊ cells. Furthermore, using small interfering RNAs to decrease ASK1 expression and a dominant negative ASK1 mutant to inhibit ASK1 kinase activity, we determined that H 2 O 2 -induced apoptosis was ASK1-dependent. Collectively, these data argue that the predisposition of Fancc ؊/؊ hematopoietic stem/progenitor cells to apoptosis is mediated in part through altered redox regulation and ASK1 hyperactivation.

Endothelial progenitor cells (EPCs) circulate in the peripheral blood and reside in blood vessel walls. A hierarchy of EPCs exists where progenitors can be discriminated based on their clonogenic potential. EPCs are exposed to oxidative stress during vascular injury as residents of blood vessel walls or as circulating cells homing to sites of neovascularization. Given the links between oxidative injury, endothelial cell dysfunction, and vascular disease, we tested whether EPCs were sensitive to oxidative stress using newly developed clonogenic assays. Strikingly, in contrast to previous reports, we demonstrate that the most proliferative EPCs (high proliferative potential-endothelial colony-forming cells and low proliferative potential-endothelial colony-forming cells) had decreased clonogenic capacity after oxidant treatment. In addition, EPCs exhibited increased apoptosis and diminished tube-forming ability in vitro and in vivo in response to oxidative stress, which was directly linked to activation of a redox-dependent stress-induced kinase pathway. Thus, this study provides novel insights into the effect of oxidative stress on EPCs. Furthermore, this report outlines a framework for understanding how oxidative injury leads to vascular disease and potentially limits the efficacy of transplantation of EPCs into ischemic tissues enriched for reactive oxygen species and oxidized metabolites.

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AUTHOR CONTRIBUTIONS D.W.C. and S.D.R. conceptualized and designed the studies evaluating cabozantinib in the preclinical NF1-mouse model. W.K.B. conducted the in vivo preclinical therapeutic studies. L.J. microdissected tumor bearing nerve tissues from experimental mice and measured proximal nerve volumes. A.E.S. performed immunohistochemical stains and conducted histopathological evaluation of all tumor specimens with assistance from S.

Objectives To determine if the spontaneous reinnervation that characteristically ensues after recurrent laryngeal nerve (RLN) injury could be selectively promoted and directed to certain laryngeal muscles with the use of neurotrophic factor (NF)-secreting muscle stem cell (MSC) vectors while antagonistic reinnervation is inhibited with vincristine (VNC). Study Design Basic science investigations involving primary cell cultures, gene cloning/transfer, and animal experiments. Methods (i.) MSC survival assays were used to test multiple individual NFs in vitro. (ii.) Motoneuron outgrowth assays assessed the trophic effects of identified NF on cranial nerve X-derived (CNX) motoneurons in vitro. (iii.) Therapeutic NF was cloned into a lentiviral vector, and MSCs were tranduced to secrete NF. 60 rats underwent left RLN transection injury, and at 3 weeks received injections of either MSCs (n=24), MSCs secreting NF (n=24), or saline (n=12) into the left thyroarytenoid muscle complex (TA); half of the animals in the MSC groups simultaneously received left posterior cricoarytenoid (PCA) injections of vincristine (VNC) while half the animals received saline. Results (i.) Ciliary-derived neurotrophic factor (CNTF) had the greatest survival-promoting effect on MSCs in culture. (ii.) Addition of CNTF (50 ng/mL) to CN X motoneuron cultures resulted in enhanced neurite outgrowth and branching. (iii.) In the animal model, the injected MSCs fused with the denervated myofibers, immunohistochemistry demonstrated enhanced reinnervation based on motor endplate to nerve contact, and RT-PCR confirmed stable CNTF expression at longest follow-up (4 months) in the CNTF-secreting MSC treated groups. Conclusions MSC therapy may have a future role in selectively promoting and directing laryngeal reinnervation after RLN injury. Level of evidence: NA

Objective Defects in the apoptotic machinery and augmented survival signals contribute to drug resistance in glioblastoma (GBM). Moreover, another complexity related to GBM treatment is the concept that GBM development and recurrence may arise from the expression of GBM stem cells (GSCs). Therefore, the use of a multifaceted approach or multitargeted agents that affect specific tumor cell characteristics will likely be necessary to successfully eradicate GBM. The objective of this study was to investigate the usefulness of sulforaphane (SFN)—a constituent of cruciferous vegetables with a multitargeted effect—as a therapeutic agent for GBM. Methods The inhibitory effects of SFN on established cell lines, early primary cultures, CD133-positive GSCs, GSC-derived spheroids, and GBM xenografts were evaluated using various methods, including GSC isolation and the sphere-forming assay, analysis of reactive oxygen species (ROS) and apoptosis, cell growth inhibition assay, comet assays for assessing SFN-triggered DNA damage, confocal microscopy, Western blot analysis, and the determination of in vivo efficacy as assessed in human GBM xenograft models. Results SFN triggered the significant inhibition of cell survival and induced apoptotic cell death, which was associated with caspase 3 and caspase 7 activation. Moreover, SFN triggered the formation of mitochondrial ROS, and SFN-triggered cell death was ROS dependent. Comet assays revealed that SFN increased single- and double-strand DNA breaks in GBM. Compared with the vehicle control cells, a significantly higher amount of γ-H2AX foci correlated with an increase in DNA double-strand breaks in the SFN-treated samples. Furthermore, SFN robustly inhibited the growth of GBM cell–induced cell death in established cell cultures and early-passage primary cultures and, most importantly, was effective in eliminating GSCs, which play a major role in drug resistance and disease recurrence. In vivo studies revealed that SFN administration at 100 mg/kg for 5-day cycles repeated for 3 weeks significantly decreased the growth of ectopic xenografts that were established from the early passage of primary cultures of GBM10. Conclusions These results suggest that SFN is a potent anti-GBM agent that targets several apoptosis and cell survival pathways and further preclinical and clinical studies may prove that SFN alone or in combination with other therapies may be potentially useful for GBM therapy.