Glioblastoma multiforme (GBM), the most common and lethal tumor of the adult brain, generally shows chemo- and radioresistance. MicroRNAs (miRs) regulate physiological processes, such as resistance of GBM cells to temozolomide (TMZ). Although miRs are attractive targets for cancer therapeutics, the effectiveness of this approach requires targeted delivery. Mesenchymal stem cells (MSCs) can migrate to the sites of cancers, including GBM. We report on an increase in miR-9 in TMZ-resistant GBM cells. miR-9 was involved in the expression of the drug efflux transporter, P-glycoprotein. To block miR-9, methods were developed with Cy5-tagged anti-miR-9. Dye-transfer studies indicated intracellular communication between GBM cells and MSCs. This occurred by gap junctional intercellular communication and the release of microvesicles. In both cases, anti-miR-9 was transferred from MSCs to GBM cells. However, the major form of transfer occurred with the microvesicles. The delivery of anti-miR-9 to the resistant GBM cells reversed the expression of the multidrug transporter and sensitized the GBM cells to TMZ, as shown by increased cell death and caspase activity. The data showed a potential role for MSCs in the functional delivery of synthetic anti-miR-9 to reverse the chemoresistance of GBM cells.

The rapid evolution of experimental data has acknowledged the critical relevance of immune biology in stem cell research. It appears that efficient transfer of stem cells to patients requires robust analyses of the immune properties as well as the responses of the stem cells to immune mediators. This review discusses the biology of adult human mesenchymal stem cells (MSCs) in the context of immunology. MSCs are pluripotent, self-renewing cells with the potential for tissue regeneration, for example the repair of bone, cartilage, tendon, ligament, skeletal muscle, and cardiac muscle. MSCs have also been shown to transdifferentiate into cells of ectodermal origin, such as neurons. MSCs are located in perfused areas of adult bone marrow, whereas hematopoietic stem cells are located in poorly perfused areas of the same organ. MSCs show bimodal, i.e. anti-inflammatory and immune-enhancing, immune responses. MSCs also regulate immune responses such as the regulation of antibody production by B cells, alterations in T cell subtypes, and immune tolerance of allogeneic transplants. MSCs also have the potential for gene delivery. This review explores the diverse clinical potential for MSCs and discusses the limitations and advantages of their immunomodulatory properties.

Glioblastoma Multiforme (GBM) is an aggressive adult primary brain tumor with poor prognosis. GBM patients develop resistance to the frontline chemotherapy, temozolomide (TMZ). As the connexins (Cx) have been shown to have a complex role in GBM, we investigated the role of Cx43 in TMZ resistance. Cx43 was increased in the TMZ-resistant low passage and cell lines. This correlated with the data in The Cancer Genome Atlas. Cx43 knockdown, reporter gene assays, chromatin immunoprecipitation assay, real-time PCR and western blots verified a role for Cx43 in TMZ resistance. This occurred by TMZ-resistant GBM cells being able to activate epidermal growth factor receptor (EGFR). In turn, EGFR activated the JNK-ERK1/2-AP-1 axis to induce Cx43. The increased Cx43 was functional as indicated by gap junctional intercellular communication among the resistant GBM cells. Cell therapy could be a potential method to deliver drugs, such as anti-EGF to tumor cells. Similar strategies could be used to reverse the expression of Cx43 to sensitize GBM cells to TMZ. The studies showed the potential for targeting EGF in immune therapy. These agents can be used in conjunction with stem cell therapy to treat GBM.

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Glioblastoma multiforme (GBM) commonly resists the frontline chemotherapy treatment temozolomide. The multidrug resistance gene (MDR1) and its protein, P-glycoprotein (P-gp), are associated with chemoresistance. This study investigated the mechanisms underlying MDR1-mediated resistance by GBM to temozolomide. P-gp trafficking was studied by flow cytometry and Western blot analysis. MDR1 expression was analyzed by real-time PCR and reporter gene assays. AP-1 interaction with MDR1 was studied by chromatin immunoprecipitation assay. EGF production was analyzed by ELISA, EGFR signaling was determined by Western blot analysis, and in vivo response to erlotinib and/or temozolomide was studied in nude mice. During the early phase of temozolomide treatment, intracellular P-gp was trafficked to the cell membrane, followed by conformational change into active P-gp. At the later phase, gene transcription of MDR1 was induced by temozolomide-mediated production of EGF. EGF activated ERK1/2-JNK-AP-1 cofactors (c-jun and c-fos). An inhibitor of EGFR kinase (erlotinib) given to nude mice with GBM prevented temozolomideinduced resistance. The results identified an essential role for activated EGFR in the resistance of GBM to temozolomide. Temozolomide resistance occurred through a biphasic response; first, by a conformational change in P-gp into the active form and, second, by releasing EGF, which caused autocrine stimulation of GBM cells to induce MDR1. Pharmacologic inhibition of EGFR kinase blunted the ability of GBM cells to resist temozolomide. These findings may explain reports on the common occurrence of mutant EGFR (EGFRvIII) and EGFR expansion in the resistance of GBM cells. Mol Cancer Ther; 13(10); 2399-411. Ó2014 AACR.

Glioblastoma Multiforme (GBM), the most common and lethal adult primary tumor of the brain, showed a link between Sonic Hedgehog (SHH) pathway in the resistance to temozolomide (TMZ). PTCH1, the SHH receptor, can tonically represses signaling by endocytosis. We asked how the decrease in PTCH1 in GBM cells could lead to TMZ-resistance. TMZ resistant GBM cells have increased PTCH1 mRNA and reduced protein. Knockdown of Dicer, a Type III RNAase, indicated that miRNAs can explain the decreased PTCH1 in TMZ resistant cells. Computational studies, real-time PCR, reporter gene studies, western blots, target protector oligos and ectopic expression identified miR-9 as the target of PTCH1 in resistant GBM cells with concomitant activation of SHH signaling. MiR-9 mediated increases in the drug efflux transporters, MDR1 and ABCG2. MiR-9 was increased in the tissues from GBM patients and in an early passage GBM cell line from a patient with recurrent GBM but not from a naïve patient. Pharmacological inhibition of SHH signaling sensitized the GBM cells to TMZ. Taken together, miR-9 targets PTCH1 in GBM cells by a SHH-independent method in GBM cells for TMZ resistance. The identified pathways could lead to new strategies to target GBM with combinations of drugs.

Several patient and surgical variables are associated with increased rate of SSI after CD. Identification of risk factors for SSI after CD is important for targeted implementation of quality improvement measures and infection control interventions.

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