David Debruyne

Subchronic alpha-linolenic acid treatment enhances brain plasticity and exerts an antidepressant effect: a versatile potential therapy for stroke.

Omega-3 polyunsaturated fatty acids are known to have therapeutic potential in several neurological and psychiatric disorders. However, the molecular mechanisms of action underlying these effects are not well elucidated. We previously showed that alpha-linolenic acid (ALA) reduced ischemic brain damage after a single treatment. To follow-up this finding, we investigated whether subchronic ALA treatment promoted neuronal plasticity. Three sequential injections with a neuroprotective dose of ALA increased neurogenesis and expression of key proteins involved in synaptic functions, namely, synaptophysin-1, VAMP-2, and SNAP-25, as well as proteins supporting glutamatergic neurotransmission, namely, V-GLUT1 and V-GLUT2. These effects were correlated with an increase in brain-derived neurotrophic factor (BDNF) protein levels, both in vitro using neural stem cells and hippocampal cultures and in vivo, after subchronic ALA treatment. Given that BDNF has antidepressant activity, this led us to test whether subchronic ALA treatment could produce antidepressant-like behavior. ALA-treated mice had significantly reduced measures of depressive-like behavior compared with vehicle-treated animals, suggesting another aspect of ALA treatment that could stimulate functional stroke recovery by potentially combining acute neuroprotection with long-term repair/compensatory plasticity. Indeed, three sequential injections of ALA enhanced protection, either as a pretreatment, wherein it reduced post-ischemic infarct volume 24 h after a 1-hour occlusion of the middle cerebral artery or as post-treatment therapy, wherein it augmented animal survival rates by threefold 10 days after ischemia.

Tumorigenic Potential of miR‐18A* in Glioma Initiating Cells Requires NOTCH‐1 Signaling

Stem cell‐like properties of glioma initiating cells (GiCs) fuel glioblastoma (GBM) development by providing the different cell types that comprise the tumor. It is therefore likely that the molecular circuitries that regulate their decision to self‐renew or commit to a more differentiated state may offer targets for future innovative therapies. In previous micro‐RNA profiling studies to search for regulators of stem cell plasticity, we identified miR‐18a* as a potential candidate and its expression correlated with the stemness state. Here, using human GiCs we found that miR‐18a* expression promotes clonal proliferation in vitro and tumorigenicity in vivo. Mechanistically, ERK‐dependent induction of miR‐18a* directly represses expression of DLL3, an autocrine inhibitor of NOTCH, thus enhancing the level of activated NOTCH‐1. Activated NOTCH‐1 in turn is required for sustained ERK activation. This feed‐forward loop, driven by miR‐18a*, is required to turn on the SHH‐GLI‐NANOG network, essential for GiC self‐renewal. Hence, by tightly regulating expression of DLL3, miR‐18a* constitutes an important signaling mediator for fine tuning the level of GiC self‐renewal. STEM Cells2013;31:1252–1265

Characterization of patient-derived tumor xenograft models of endometrial cancer for preclinical evaluation of targeted therapies.

OBJECTIVE Endometrial carcinoma (EC) is the sixth most common cancer in women and therapies are limited for advanced and recurrent disease. Patient-derived tumor xenograft (PDTX) models are becoming popular tools in translational research because of their histological and genetic similarity to the original tumors and the ability to predict therapeutic response to treatments. Here, we established and characterized a panel of 24 EC PDTX models which includes the major histological and genetic subtypes observed in patients. METHODS Fresh tumor tissues collected from primary, metastatic and recurrent type I and type II EC patients were engrafted in immunocompromised mice. Histology, vimentin, and cytokeratin expression were evaluated, together with Microsatellite instability (MSI), mutation profiling by Whole Exome Sequencing and copy number profiling by Whole Genome Low Coverage Sequencing. The efficacy of both PI3K and MEK inhibitors was evaluated in a model of endometrioid carcinoma harboring PTEN, PIK3CA and KRAS mutations. RESULTS We observed good similarity between primary tumors and the corresponding xenografts, at histological and genetic level. Among the engrafted endometrioid models, we found a significant enrichment of MSI and POLE mutated tumors, compared to non-engrafted samples. Combination treatment with NVP-BEZ235 and AZD6244 showed the possibility to stabilize the tumor growth in one model originated from a patient who already received several lines of chemotherapy. CONCLUSION The established EC PDTX models, resembling the original human tumors, promise to be useful for preclinical evaluation of novel combination and targeted therapies in specific EC subgroups.

The pH-dependent photoluminescence of colloidal CdSe/ZnS quantum dots with different organic coatings.

The photoluminescence (PL) of colloidal quantum dots (QDs) is known to be sensitive to the solution pH. In this work we investigate the role played by the organic coating in determining the pH-dependent PL. We compare two types of CdSe/ZnS QDs equipped with different organic coatings, namely dihydrolipoic acid (DHLA)-capped QDs and phospholipid micelle-encapsulated QDs. Both QD types have their PL intensity quenched at acidic pH values, but they differ in terms of the reversibility of the quenching process. For DHLA-capped QDs, the quenching is nearly irreversible, with a small reversible component visible only on short time scales. For phospholipid micelle-encapsulated QDs the quenching is notably almost fully reversible. We suggest that the surface passivation by the organic ligands is reversible for the micelle-encapsulated QDs. Additionally, both coatings display pH-dependent spectral shifts. These shifts can be explained by a combination of irreversible processes, such as photo-oxidation and acid etching, and reversible charging of the QD surface, leading to the quantum-confined Stark effect (QCSE), the extent of each effect being coating-dependent. At high ionic strengths, the aggregation of QDs also leads to a spectral (red) shift, which is attributable to the QCSE and/or electronic energy transfer.

Validation of the Freund Clock Drawing Test as a screening tool to detect cognitive dysfunction in elderly cancer patients undergoing comprehensive geriatric assessment.

We aimed to validate the Freund Clock Drawing Test (CDT), with its predefined cutoff score of ≤4, as a screening tool to detect elderly cancer patients in need of a more in‐depth cognitive evaluation within a comprehensive geriatric assessment (CGA).

Microsatellite instable and microsatellite stable primary endometrial carcinoma cells and their subcutaneous and orthotopic xenografts recapitulate the characteristics of the corresponding primary tumor

Objective Well-characterized, low-passage, primary cell cultures established directly from patient tumors are an important tool for drug screening because these cultures faithfully recapitulate the genomic features of primary tumors. Here, we aimed to establish these cell cultures from primary endometrial carcinomas (ECs) and to develop subcutaneous and orthotopic xenograft models as a model to validate promising treatment options for EC in the in vivo setting. Methods Primary cell cultures of EC tumors were established and validated by analysing histologic and genetic characteristics, telomerase activity, and in vitro and in vivo growth characteristics. Using these primary cell cultures, subcutaneous and orthotopic mouse models were subsequently established. Results We established and characterized 7 primary EC cell cultures and corresponding xenograft models of different types of endometrioid tumors. Interestingly, we observed that the chance to successfully establish a primary cell culture seems higher for microsatellite instable than microsatellite stable tumors. For the first time, we also established an orthotopic murine model for EC derived from a primary cell culture. In contrast to EC cell lines, grafted tumor cultures preserved the original tumor structure and mimicked all histologic features. They also established abdominal and distant metastases, reflecting the tumorigenic behavior in the clinical setting. Remarkably, the established cell cultures and xenograft tumors also preserved the genetic characteristics of the primary tumor. Conclusions The established EC cultures reflect the epithelial genetic characteristics of the primary tumor. Therefore, they provide an appropriate model to investigate EC biology and apply high-throughput drug screening experiments. In addition, the established murine xenograft models, in particular the orthotopic model, will be useful to validate promising therapeutic strategies in vivo, as the grafted tumors closely resemble the primary tumors from which they were derived. Microsatellite instable status seems to determine the success rate of establishing primary cell cultures.

Enzyme conjugation and biosensing with quantum dots: A photoluminescence study

In this paper we investigate the enzyme conjugation of CdSe/ZnS core/shell quantum dots (QDs) by means of their photoluminescent properties, as well as the influence of the enzymatic reaction on these properties. The QDs were first transferred from toluene to water by encapsulation in poly(ethylene glycol)-phospholipid micelles. The water solubilized QDs were bioconjugated with the enzymes glucose oxidase (GOX) and horseradish peroxidase (HRP). For covalent coupling the crosslinkers 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and hydroxysulfosuccinimide (sulfo-NHS) were used. Photoluminescence (PL) measurements showed that EDC had a large effect on the PL intensity. Also, the PL intensity increased in PBS buffer as compared to water. The enzyme conjugation caused a noticeable shift in the PL peak wavelength, but had a minor effect on the PL intensity. Sensing experiments confirmed the well-known quenching of PL by H2O2. Similarly, the PL intensity was sensitive to glucose in the mM range in the presence of GOX. This can be understood since the conversion of glucose by GOX yields H2O2 as a by-product. Measurements of the quantum yield (QY) showed increased values for QD solutions treated with EDC. The QY decreased with increasing glucose concentration in the presence of GOX. The change in QY was accompanied by a change in pH, suggesting a correlation between pH and QY.

Establishment and characterization of uterine sarcoma and carcinosarcoma patient-derived xenograft models

OBJECTIVE Uterine sarcomas (US) and carcinosarcomas (CS) are rare, aggressive cancers. The lack of reliable preclinical models hampers the search for new treatment strategies and predictive biomarkers. To this end, we established and characterized US and CS patient-derived xenograft (PDX) models. METHODS Tumor fragments of US and CS were subcutaneously implanted into immunocompromised mice. Engrafted xenograft and original tumors were compared by means of histology, immunohistochemistry, whole-genome low-coverage sequencing for copy number variations, and RNA sequencing. RESULTS Of 13 implanted leiomyosarcomas (LMS), 10 engrafted (engraftment rate of 77%). Also 2 out of 7 CS (29%) and one high-grade US (not otherwise specified) models were successfully established. LMS xenografts showed high histological similarity to their corresponding human tumors. Expression of desmin and/or H-caldesmon was detected in 8/10 LMS PDX models. We noticed that in CS models, characterized by the concomitant presence of a mesenchymal and an epithelial component, the relative distribution of the components is varying over the generations, as confirmed by changes in vimentin and cytokeratin expression. The similarity in copy number profiles between original and xenograft tumors ranged from 57.7% to 98.2% for LMS models and from 47.4 to 65.8% for CS models. Expression pattern stability was assessed by clustering RNA expression levels of original and xenograft tumors. Six xenografts clustered together with their original tumor, while 3 (all LMS) clustered apart. CONCLUSIONS We present here a panel of clinically annotated uterine sarcoma and carcinosarcoma PDX models, which will be a useful tool for preclinical testing of new therapies.