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Several pharmacological approaches are used for glioblastoma (GBM) treatment, each hinging on the triggering of different biochemical or functional processes; the development of specific and sensitive PET procedures for monitoring their efficacy proceeds with the identification of such new treatments. This paper presents an overview of the available “tumour biomarker”–“PET probe” pairs (i.e. the combination of a tumour target and a selective PET radiopharmaceutical) for monitoring the different treatments for GBM tested in human subjects. A bibliographic search for papers on PET imaging for assessing treatment response in GBM was performed in PubMed and Web of science databases using the following string: (PET or positron) and (glioblastoma) and (treatment) and (monitoring); papers dealing with studies in human subjects published over the last 10 years were reviewed. Further papers were extracted from the bibliography of the reviewed papers. In this review, we highlight through a detailed table that in spite of the current use in GBM patients of a large variety of PET radiopharmaceuticals, very few papers have specifically addressed the issue of the optimization and use of imaging biomarker–probe pairs for the assessment of treatment response in GBM. While new PET probes are being developed for assessing old and new GBM biomarkers, very few clinical trials have been performed to this end. Whereas it appears that the use of old and new PET radiopharmaceuticals can advance the non-invasive assessment of treatment response in GBM, the optimal match of biomarker–probe pairs although highly needed is still being sought in particular with the active development of new highly specific treatments characterized by novel antitumoral targeting strategies.

Chaperone-mediated autophagy (CMA) is a catabolic pathway fundamental for cell homeostasis, by which specific damaged or non-essential proteins are degraded. CMA activity has three main levels of regulation. The first regulatory level is based on the targetability of specific proteins possessing a KFERQ-like domain, which can be recognized by specific chaperones and delivered to the lysosomes. Target protein unfolding and translocation into the lysosomal lumen constitutes the second level of CMA regulation and is based on the modulation of Lamp2A multimerization. Finally, the activity of some accessory proteins represents the third regulatory level of CMA activity. CMA's role in oncology has not been fully clarified covering both pro-survival and pro-death roles in different contexts. Taking all this into account, it is possible to comprehend the actual complexity of both CMA regulation and the cellular consequences of its activity allowing it to be elected as a modulatory and not only catabolic machinery. In this review, the role covered by CMA in oncology is discussed with a focus on its relevance in glioma. Molecular correlates of CMA importance in glioma responsiveness to treatment are described to identify new early efficacy biomarkers and new therapeutic targets to overcome resistance.

Glioblastoma (GBM) is the most aggressive tumor of CNS characterized by a poor prognosis mainly due to the resistance to the available treatments. Temozolomide (TMZ) is the gold standard for GBM treatment although this drug even in association with radiotherapy provides a median survival of 14.6 months. There are evidences that the increase in intracellular ROS is able to sustain gliomagenesis, tumor proliferation rate, and resistance to treatments by supporting proliferative pathways and metabolism as well as by generating genetic instability causing the establishment of protumorigenic mutants. At the same time, an increase in intracellular ROS, being involved in the oxidative damage of DNA, lipids, and proteins, is involved in the impairment of cell function, triggering also cell death. Reactive oxygen species (ROS) are highly reactive molecules originating from different enzymatic and nonenzymatic reactions occurring in mitochondria and other different intracellular organelles. They can have also an exogenous origin related to the effects of xenobiotics or radiations. Under basal conditions, ROS generation and their scavenging are tightly controlled, and redox homeostasis is maintained, but in case of an imbalance in ROS turnover, oxidative stress can occur producing cell damage. For these reasons, ROS modulation has been pursued by different treatment plans to increase cytotoxicity and overcome GBM resistance to treatment. Since the double-edged sword role covered by ROS, which mediates cell survival signaling, both a positive and a negative ROS modulation have been assessed for therapeutic purposes. In this chapter, the mechanisms involved in the relation between ROS and cell response to treatment are described with particular attention focused on the induction of chaperone-mediated autophagy (CMA) whose activity together with the reduction of hypoxia-inducible factor-1α (HIF-1) activity after treatment have been related to GBM cell responsiveness to TMZ.

Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with poor survival. Cytoreduction in association with radiotherapy and temozolomide (TMZ) is the standard therapy, but response is heterogeneous and life expectancy is limited. The combined use of chemotherapeutic agents with drugs targeting cell metabolism is becoming an interesting therapeutic option for cancer treatment. Here, we found that metformin (MET) enhances TMZ effect on TMZ-sensitive cell line (U251) and overcomes TMZ-resistance in T98G GBM cell line. In particular, combined-treatment modulated apoptosis by increasing Bax/Bcl-2 ratio, and reduced Reactive Oxygen Species (ROS) production. We also observed that MET associated with TMZ was able to reduce the expression of glioma stem cells (GSC) marker CD90 particularly in T98G cells but not that of CD133. In vivo experiments showed that combined treatment with TMZ and MET significantly slowed down growth of TMZ-resistant tumors but did not affect overall survival of TMZ-sensitive tumor bearing mice. In conclusion, our results showed that metformin is able to enhance TMZ effect in TMZ-resistant cell line suggesting its potential use in TMZ refractory GBM patients. However, the lack of effect on a GBM malignancy marker like CD133 requires further evaluation since it might influence response duration.

Hypoxia is a common feature in solid tumours. In glioma, it is considered the major driving force for tumour angiogenesis and correlates with enhanced resistance to conventional therapies, increased invasiveness and a poor prognosis for patients. Here we describe, for the first time, that miR675-5p, embedded in hypoxia-induced long non-coding RNA H19, plays a mandatory role in establishing a hypoxic response and in promoting hypoxia-mediated angiogenesis. We demonstrated, in vitro and in vivo, that miR675-5p over expression in normoxia is sufficient to induce a hypoxic moreover, miR675-5p depletion in low oxygen conditions, drastically abolishes hypoxic responses including angiogenesis. In addition, our data indicate an interaction of miR675-5p, HIF-1α mRNA and the RNA Binding Protein HuR in hypoxia-induced responses. We suggest the modulation of miR675-5p as a new therapeutic option to promote or abolish hypoxia induced angiogenesis.

Interaction between tumor cells and the microenvironment is key in initiation, progression, and invasiveness of cancer. In particular, mesenchymal stem cells (MSCs) are recruited to the sites of developing tumors, thus promoting metastasis formation. Although it is well known that MSCs migrate and integrate in the tumor microenvironment (TME), their fate and function inside the tumor is still not clear. In this study, we analyzed the role played by MSCs in breast cancer oncogenesis. Data indicate that interaction of breast cancer cells with MSCs results in an increased proliferation and metabolic activity of breast cancer cells, partially due to MSC-derived microvesicles that are shed in the TME. Moreover, we addressed the question of whether we could modulate such interaction by acting on P2X-mediated intercellular communication. By inhibiting P2X-mediated purinergic signaling, we succeeded in reducing both the cancerogenic as well as the metastatic potential of breast cancer cells co-cultured with MSCs, in 2D as well as in 3D in vitro models. Data obtained demonstrate for the first time that the trophic effect of MSCs on breast cancer cell growth is exerted via ionotropic purinergic signaling, thus suggesting the inhibition of the purinergic signaling system as a potential target for therapeutic intervention.

Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with poor survival. Cytoreduction in association with radiotherapy and temozolomide (TMZ) is the standard therapy, but response is heterogeneous and life expectancy is limited. The combined use of chemotherapeutic agents with drugs targeting cell metabolism is becoming an interesting therapeutic option for cancer treatment. Here, we found that metformin (MET) enhances TMZ effect on TMZ-sensitive cell line (U251) and overcomes TMZ-resistance in T98G GBM cell line. In particular, combined-treatment modulated apoptosis by increasing Bax/Bcl-2 ratio, and reduced Reactive Oxygen Species (ROS) production. We also observed that MET associated with TMZ was able to reduce the expression of glioma stem cells (GSC) marker CD90 particularly in T98G cells but not that of CD133. In vivo experiments showed that combined treatment with TMZ and MET significantly slowed down growth of TMZ-resistant tumors but did not affect overall survival of TMZ-sensitive tumor bearing mice. In conclusion, our results showed that metformin is able to enhance TMZ effect in TMZ-resistant cell line suggesting its potential use in TMZ refractory GBM patients. However, the lack of effect on a GBM malignancy marker like CD133 requires further evaluation since it might influence response duration.

Cancer is a complex disease, characterized by alteration of different physiological molecular processes and cellular features. Keeping this in mind, the possibility of early identification and detection of specific tumor biomarkers by non-invasive approaches could improve early diagnosis and patient management. Different molecular imaging procedures provide powerful tools for detection and non-invasive characterization of oncological lesions. Clinical studies are mainly based on the use of computed tomography, nuclear-based imaging techniques and magnetic resonance imaging. Preclinical imaging in small animal models entails the use of dedicated instruments, and beyond the already cited imaging techniques, it includes also optical imaging studies. Optical imaging strategies are based on the use of luminescent or fluorescent reporter genes or injectable fluorescent or luminescent probes that provide the possibility to study tumor features even by means of fluorescence and luminescence imaging. Currently, most of these probes are used only in animal models, but the possibility of applying some of them also in the clinics is under evaluation. The importance of tumor imaging, the ease of use of optical imaging instruments, the commercial availability of a wide range of probes as well as the continuous description of newly developed probes, demonstrate the significance of these applications. The aim of this review is providing a complete description of the possible optical imaging procedures available for the non-invasive assessment of tumor features in oncological murine models. In particular, the characteristics of both commercially available and newly developed probes will be outlined and discussed.

INTRODUCTION: Gliomas represent the most common adult neoplasm of the primary central nervous system (CNS) characterized by wide necrotic areas, high invasiveness, resistance to therapy and poor prognosis. Glioma is indeed refractory to conventional treatment combining surgery, radiotherapy and chemiotherapy (mostly Temozolomide, TMZ). Hypoxia is considered a major driving force for glioma progression inducing chemio- and radio-resistance, and sustaining tumor angiogenesis and invasiveness as well as proliferation with the hypoxia-mediated increase of HIF-1α activity. The presented research is part of the INSERT (INtegrated SPECT/MRI for Enhanced Stratification in Radio-chemo Therapy) project, a new collaborative research project founded by the European Seventh Research Framework Program (FP7) aimed at the development of a multi-modality imaging tool for concurrent SPECT and MRI studies to provide improved personalized radio-chemo therapies for brain tumors. Within this frame, the aim of the work, described herein, has been the identification of non-invasive theranostic biomarkers, involved in the regulation of tumor growth, and for the early assessment of tumor response to therapeutics using in vivo imaging strategies in an orthotopic mouse model of glioma. METHODS: Murine model was obtained by stereotaxic injection of human glioma cells (U251-HRE expressing Luciferase reporter gene under the control of multiple copies of hypoxia responsive element). Mice were treated with TMZ 400 mg/kg o.s 21 days after cell injection. Tumor growth was monitored in vivo with MRI (7 Tesla Bruker scanner) and Optical imaging (BLI and FLI, IVIS SPECTRUM/CT instrument), before and after treatment (0, 2 and 7 days). Moreover, ex vivo preliminary uptake studies were performed by autoradiography with [99mTc]MIBI. For MRI examinations were used different sequences such as: EPI-DTI (Diffusion Tensor), Turbo Spin Echo T2, Rare T1 Map, Dynamic Contrast Enhancement (DCE) Dynamic Susceptibility Contrast (DSC) and T1 post-contrast. RESULTS: TMZ treatment, beyond the already published reduction of HIF-1 activity, induced a reduction of tumor volume reported both by MRI and autoradiography one week after treatment. Treatment efficacy induced also variation of MRI detectable phenomena, such as diffusion and contrast enhancement (DWI and DCE MRI sequences). [99mTc]MIBI uptake resulted significantly lowered after TMZ treatment allowing the identification of responsive tumors. These data correlate with those already obtained by BLI and FLI. CONCLUSIONS: This study suggests that, in the U251-HRE orthotopic murine model, the responsiveness to TMZ treatment induced a pattern of changes probably involved with HIF-1 activity. These changes can be monitored by multiparametric analysis, and will be assessed using the newly hybrid imaging system proposed by INSERT project, allowing the validation of the new theranostic biomarkers of early tumor response to treatment. ACKNOWLEDGMENT: This work was supported by the FP7 INSERT project (HEALTH-2012-INNOVATION-1, GA305311).

Background/Aims: Congenital hyperinsulinism of infancy is a rare disease that needs prompt treatment to avoid brain damage. There are currently no data regarding the clinical and molecular features of Italian patients. Methods: Thirty-three patients with HI and their parents were included. Consanguinity was reported in six patients. Half of patients were macrosomic at birth. None had raised 3-hydroxybutyrylcarnitine or hyperammonemia. Molecular analysis of ABCC8 and KCNJ11 genes was performed in all patients, and subjects with no mutation underwent analysis of HNF4A and GCK. GLUD1 and HADH genes were analyzed in a patient with leucine sensitivity. Results: Mutations in the ABCC8 and KCNJ11 genes were found in 45% of the patients (6 novel). No mutations in HNF4A, GLUD1 and GCK genes were found. Recessive mode of inheritance was found in 21% of patients. A single heterozygous mutation was identified in 24% of probands. 72% of the patients were responsive to medical treatment, and 44% of the 17 patients with no identified mutation achieved spontaneous remission. Nine children, unresponsive to medical therapy, underwent pancreatectomy. Conclusion: This is the first report on hyperinsulinism of infancy in Italy, confirming the complexity of the clinical forms and the heterogeneity of the genetic causes of the disease.

Treatment with antiretroviral agents (ARVs) during pregnancy is important to prevent mother-to-child transmission of the human immunodeficiency virus (HIV), but their use has been associated with low bone mineral density in adult patients. Currently, there are no data regarding the bone status of HIV-infected women who received ARV during pregnancy. The aim of this study was to evaluate cortical bone status at delivery in a group of HIV-infected women who received ARV during pregnancy and to monitor the changes occurring during the first year postpartum. We studied 33 HIV-infected and 116 HIV-uninfected healthy Caucasian women within 4 days from delivery. Follow-up measurements were performed at 4 and 12 months postpartum in 17 HIV-infected and 55 healthy women. Cortical bone status was evaluated by quantitative ultrasonography at the mid-tibia, and bone measurements were expressed as the speed of sound (SOS). HIV-infected women after delivery had a median SOS of 3,985 (3,567–4,242) m/s, while the median SOS of healthy women was 4,025 (3,643–4,250) m/s. The difference was not significant (t = 0.39, P = 0.69). No significant differences were observed between ARV-exposed and control subjects at 4 and 12 months. Our data suggest that ARV during pregnancy and the first year after delivery does not affect negatively cortical bone status.