The Temozolomide-Doxorubicin paradox in Glioblastoma in vitro-in silico preclinical drug-screening.

Adjuvant Temozolomide is considered the front-line Glioblastoma chemotherapeutic treatment; yet not all patients respond. Latest trends in clinical trials usually refer to Doxorubicin; yet it can lead to severe side-effects if administered in high doses. While Glioblastoma prognosis remains poor, little is known about the combination of the two chemotherapeutics. Patient-derived spheroids were generated and treated with a range of Temozolomide/Doxorubicin concentrations either as monotherapy or in combination. Optical microscopy was used to monitor the growth pattern and cell death. Based on the monotherapy experiments, we developed a probabilistic mathematical framework in order to describe the drug-induced effect at the single-cell level and simulate drug doses in combination assuming probabilistic independence. Doxorubicin was found to be effective in doses even four orders of magnitude less than Temozolomide in monotherapy. The combination therapy doses tested in vitro were able to lead to irreversible growth inhibition at doses where monotherapy resulted in relapse. In our simulations, we assumed both drugs are anti-mitotic; Temozolomide has a growth-arrest effect, while Doxorubicin is able to cumulatively cause necrosis. Interestingly, under no mechanistic synergy assumption, the in silico predictions underestimate the in vitro results. In silico models allow the exploration of a variety of potential underlying hypotheses. The simulated-biological discrepancy at certain doses indicates a supra-additive response when both drugs are combined. Our results suggest a Temozolomide-Doxorubicin dual chemotherapeutic scheme to both disable proliferation and increase cytotoxicity against Glioblastoma.

Transrectal Absorber Guide Raster-Scanning Optoacoustic Mesoscopy for Label-Free In Vivo Assessment of Colitis.

Optoacoustic imaging (OAI) enables microscale imaging of endogenous chromophores such as hemoglobin at significantly higher penetration depths compared to other optical imaging technologies. Raster-scanning optoacoustic mesoscopy (RSOM) has recently been shown to identify superficial microvascular changes associated with human skin pathologies. In animal models, the imaging depth afforded by RSOM can enable entirely new capabilities for noninvasive imaging of vascular structures in the gastrointestinal tract, but exact localization of intra-abdominal organs is still elusive. Herein the development and application of a novel transrectal absorber guide for RSOM (TAG-RSOM) is presented to enable accurate transabdominal localization and assessment of colonic vascular networks in vivo. The potential of TAG-RSOM is demonstrated through application during mild and severe acute colitis in mice. TAG-RSOM enables visualization of transmural vascular networks, with changes in colon wall thickness, blood volume, and OAI signal intensities corresponding to colitis-associated inflammatory changes. These findings suggest TAG-RSOM can provide a novel monitoring tool in preclinical IBD models, refining animal procedures and underlines the capabilities of such technologies to address inflammatory bowel diseases in humans.

Phenocopying Glioblastoma: A Review.

The main reason why therapeutic schemes fail in Glioblastoma lies on its own peculiarities as a cancer and on our failure to fully decipher them. Fast tumor evolution, invasiveness and incomplete surgical resection contribute to disease recurrence, therapy resistance and high mortality. More faithful models must be developed to address Glioblastoma biology and better clinical guidance. Research studies are discussed in this review that: i) improve understanding and assessment of the growth mechanisms of Glioblastoma and ii) develop preclinical models (in vitro-in vivo-in silico) that mimic patient's tumor (phenocopying) in order to provide better prediction of response to therapies.

The Potential of Photoacoustic Imaging in Radiation Oncology.

Radiotherapy is recognized globally as a mainstay of treatment in most solid tumors and is essential in both curative and palliative settings. Ionizing radiation is frequently combined with surgery, either preoperatively or postoperatively, and with systemic chemotherapy. Recent advances in imaging have enabled precise targeting of solid lesions yet substantial intratumoral heterogeneity means that treatment planning and monitoring remains a clinical challenge as therapy response can take weeks to manifest on conventional imaging and early indications of progression can be misleading. Photoacoustic imaging (PAI) is an emerging modality for molecular imaging of cancer, enabling non-invasive assessment of endogenous tissue chromophores with optical contrast at unprecedented spatio-temporal resolution. Preclinical studies in mouse models have shown that PAI could be used to assess response to radiotherapy and chemoradiotherapy based on changes in the tumor vascular architecture and blood oxygen saturation, which are closely linked to tumor hypoxia. Given the strong relationship between hypoxia and radio-resistance, PAI assessment of the tumor microenvironment has the potential to be applied longitudinally during radiotherapy to detect resistance at much earlier time-points than currently achieved by size measurements and tailor treatments based on tumor oxygen availability and vascular heterogeneity. Here, we review the current state-of-the-art in PAI in the context of radiotherapy research. Based on these studies, we identify promising applications of PAI in radiation oncology and discuss the future potential and outstanding challenges in the development of translational PAI biomarkers of early response to radiotherapy.

PML Differentially Regulates Growth and Invasion in Brain Cancer.

Glioblastoma is the most malignant brain tumor among adults. Despite multimodality treatment, it remains incurable, mainly because of its extensive heterogeneity and infiltration in the brain parenchyma. Recent evidence indicates dysregulation of the expression of the Promyelocytic Leukemia Protein (PML) in primary Glioblastoma samples. PML is implicated in various ways in cancer biology. In the brain, PML participates in the physiological migration of the neural progenitor cells, which have been hypothesized to serve as the cell of origin of Glioblastoma. The role of PML in Glioblastoma progression has recently gained attention due to its controversial effects in overall Glioblastoma evolution. In this work, we studied the role of PML in Glioblastoma pathophysiology using the U87MG cell line. We genetically modified the cells to conditionally overexpress the PML isoform IV and we focused on its dual role in tumor growth and invasive capacity. Furthermore, we targeted a PML action mediator, the Enhancer of Zeste Homolog 2 (EZH2), via the inhibitory drug DZNeP. We present a combined in vitro-in silico approach, that utilizes both 2D and 3D cultures and cancer-predictive computational algorithms, in order to differentiate and interpret the observed biological results. Our overall findings indicate that PML regulates growth and invasion through distinct cellular mechanisms. In particular, PML overexpression suppresses cell proliferation, while it maintains the invasive capacity of the U87MG Glioblastoma cells and, upon inhibition of the PML-EZH2 pathway, the invasion is drastically eliminated. Our in silico simulations suggest that the underlying mechanism of PML-driven Glioblastoma physiology regulates invasion by differential modulation of the cell-to-cell adhesive and diffusive capacity of the cells. Elucidating further the role of PML in Glioblastoma biology could set PML as a potential molecular biomarker of the tumor progression and its mediated pathway as a therapeutic target, aiming at inhibiting cell growth and potentially clonal evolution regarding their proliferative and/or invasive phenotype within the heterogeneous tumor mass.

A 3D tumor spheroid model for the T98G Glioblastoma cell line phenotypic characterization.

Major Glioblastoma's hallmarks include proliferation, invasion and heterogeneity. Biological 3D tumor spheroid models can serve as intermediate systems between traditional 2D cell culture and complex in vivo models. Tumor spheroids have been shown to more accurately reproduce the spatial organization and microenvironmental factors of in vivo micro-tumors, such as relevant gradients of nutrients and other molecular agents, while they maintain cell-to-cell and cell-to-matrix interactions. In vitro 3D assays are useful to monitor these properties. Here, we test the suitability of the well-known T98 G Glioblastoma cell line in such a 3D assay. The doubling time and death rate parameters of T98 G are estimated, as well as their spheroidal growth-expansion curves with and without the presence of basement membrane substrate. The T98 G invasive profile is characterized by collective morphology and proliferation-associated invasion. We show that the T98 G secondary GB cell line exhibits both invasive and proliferative capabilities in 3D and thus, can serve as control cell line for the 3D in vitro study of primary GB cell cultures.

High resolution volumetric imaging of primary and secondary tumor spheroids using multi-angle Light Sheet Fluorescence Microscopy (LSFM).

Breast cancer and Glioblastoma brain cancer are severe malignancies with poor prognosis. In this study, primary Glioblastoma and secondary breast cancer spheroids are formed and treated with the well-known Temozolomide and Doxorubicin chemotherapeutics, respectively. High resolution imaging of both primary and secondary cancer cell spheroids is possible using a custom multi-angle Light Sheet Fluorescence Microscope. Such a technique is successful in realizing preclinical drug screening, while enables the discrimination among physiologic tumor parameters.

(1)H-NMR based metabolomics study for the detection of the human urine metabolic profile effects of Origanum dictamnus tea ingestion.

(1)H NMR spectroscopy was employed to investigate the repercussion of Origanum dictamnus tea ingestion in several volunteers' urine metabolic profiles, among them two with chronic inflammatory bowel diseases (IBD), mild IBD and Crohn's disease. Herein, we demonstrate that the concentrations of a lot of urinary metabolites such as hippurate, trimethylamine oxide (TMAO), citrate, and creatinine are altered, which prompts the intestinal microflora function/content perturbation as well as kidney function regulation by dictamnus tea. Interestingly, our preliminary results showed that a high dose of dictamnus tea intake appeared to be toxic for a person with Crohn's disease, since it caused high endogenous ethanol excretion in urine. All subjects' metabolic effects caused by the dictamnus tea appeared to be reversible, when all volunteers stopped its consumption. Finally, we highlight that individuals' metabolic phenotype is reflected in their urine biofluid before and after the dictamnus tea effect while all individuals have some common and different metabolic responses to this tea, implying that each phenotype has a quite different response to this tea consumption.

A Proposed Paradigm Shift in Initializing Cancer Predictive Models with DCE-MRI Based PK Parameters: A Feasibility Study.

Glioblastoma multiforme is the most aggressive type of glioma and the most common malignant primary intra-axial brain tumor. In an effort to predict the evolution of the disease and optimize therapeutical decisions, several models have been proposed for simulating the growth pattern of glioma. One of the latest models incorporates cell proliferation and invasion, angiogenic net rates, oxygen consumption, and vasculature. These factors, particularly oxygenation levels, are considered fundamental factors of tumor heterogeneity and compartmentalization. This paper focuses on the initialization of the cancer cell populations and vasculature based on imaging examinations of the patient and presents a feasibility study on vasculature prediction over time. To this end, pharmacokinetic parameters derived from dynamic contrast-enhanced magnetic resonance imaging using Toft's model are used in order to feed the model. K (trans) is used as a metric of the density of endothelial cells (vasculature); at the same time, it also helps to discriminate distinct image areas of interest, under a set of assumptions. Feasibility results of applying the model to a real clinical case are presented, including a study on the effect of certain parameters on the pattern of the simulated tumor.