SCREENER, an educational game for teaching the Drug Discovery and Development process.

Although the use of games as an educational strategy is an important current trend, there is practically no option available for training people on the Drug Discovery and Development (DDD) process. To fill this gap, we designed "SCREENER", a science game that is intended to be educational, but also challenging and interesting enough to ensure player engagement. Our main target audience is students of postgraduate programs in pharmacology, medicinal chemistry, pharmacy, and medicine. This game could also be of interest to the pharmaceutical industry and regulatory and patent agencies for training new employees. We discuss the creation of SCREENER, a hybrid of board and card games, and present its components with some examples of cards and resources, as well as the dynamics of the game. SCREENER mimics the process of drug discovery and development from validating a target to registering the new drug with the regulatory agency, and can be played individually (self-learning) or with the help of a monitor who assists up to six players/teams. Briefly, 29 task cards categorized in four major areas (efficacy, safety, pharmacokinetics, and pharmaceutical development) must be purchased sequentially. Classic characteristics of games such as decision making and challenge have been incorporated. More in-depth information on the tasks and technical terms is available through QR codes. The vagaries of the DDD process are mimicked by the bonus/setback cards. The evaluation of our first test with students is presented and supports the usefulness of this new tool.

SCREENER, an educational game for teaching the Drug Discovery and Development process

Although the use of games as an educational strategy is an important current trend, there is practically no option available for training people on the Drug Discovery and Development (DDD) process. To fill this gap, we designed "SCREENER", a science game that is intended to be educational, but also challenging and interesting enough to ensure player engagement. Our main target audience is students of postgraduate programs in pharmacology, medicinal chemistry, pharmacy, and medicine. This game could also be of interest to the pharmaceutical industry and regulatory and patent agencies for training new employees. We discuss the creation of SCREENER, a hybrid of board and card games, and present its components with some examples of cards and resources, as well as the dynamics of the game. SCREENER mimics the process of drug discovery and development from validating a target to registering the new drug with the regulatory agency, and can be played individually (self-learning) or with the help of a monitor who assists up to six players/teams. Briefly, 29 task cards categorized in four major areas (efficacy, safety, pharmacokinetics, and pharmaceutical development) must be purchased sequentially. Classic characteristics of games such as decision making and challenge have been incorporated. More in-depth information on the tasks and technical terms is available through QR codes. The vagaries of the DDD process are mimicked by the bonus/setback cards. The evaluation of our first test with students is presented and supports the usefulness of this new tool.

Intrathecal transplantation of stem cells by lumbar puncture for thoracic spinal cord injury in the rat.

STUDY DESIGN: Experimental investigation of intrathecal transplantation of stem cells by lumbar puncture (LP) in a rat model that simulates human thoracic spinal cord injury (SCI). OBJECTIVES: To examine the distribution and phenotype of spinal cord-derived neural stem/progenitor cells (NSPCs) and bone marrow-derived mesenchymal stromal cells (BMSCs) following LP transplantation in SCI rats. SETTING: Toronto Western Research Institute, Toronto, Ontario, Canada. METHODS: NSPCs or BMSCs were transplanted via LP at level L3-5 1 week after compression SCI at T8. Rats were killed at 3, 17 and 27 days after LP transplantation and the relative distribution of cells at C4, T8 and L3-5 was quantitated. The phenotype of the NSPC and BMSC was assessed with immunocytochemistry in vitro and following LP transplantation. RESULTS: By 4 weeks, more NSPC migrated to the lesion site relative to BMSC and uninjured animals. However, there was no preferential homing of either of these types of cells into the parenchyma of the injury site, and most of the transplanted cells remained in the intrathecal space. In vitro, spinal cord-derived NSPC proliferated and expressed nestin, but after LP transplantation, NSPC became post-mitotic and primarily expressed oligodendrocyte markers. In contrast, BMSC did not express any neural antigens in vivo. CONCLUSION: LP is a minimally invasive method of cell transplantation that produces wide dissemination of cells in the subarachnoid space of the spinal cord. This is the first study to report and quantify the phenotype and spatial distribution of LP transplanted NSPC and BMSC in the intact and injured spinal cord.

Proliferation, migration, and differentiation of endogenous ependymal region stem/progenitor cells following minimal spinal cord injury in the adult rat.

Ependymal cells of the adult mammalian spinal cord exhibit stem/progenitor cell properties following injury. In the present study, we utilized intraventricular injection of 1,1'-dioctadecyl-6,6'-di(4-sulfophenyl)-3,3,3',3'-tetramethylindocarbocyanine (DiI) to label the ependyma lining the central canal to allow tracking of the migration of endogenous ependymal cells and their progeny after spinal cord injury (SCI). We developed a minimal injury model that preserved the integrity of the central canal and did not interfere with ependymal cell labeling. Three days following SCI, there was an 8.6-fold increase in the proliferative labeling index of the ependymal cells at the level of the needle track based on bromodeoxyuridine labeling, compared with 1 day post-injury. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) positive cells were not detected in the ependyma or surrounding gray matter, indicating that ependymal cells do not undergo apoptosis in response to minimal injury. Nestin was rapidly induced in the ependyma by 1 day and expression peaked by 7 days post-injury. We quantitated the number and distance of ependymal cell migration following minimal injury. The number of ependymal cells migrating from the region of the central canal increased by 3 days following minimal injury and DiI-labeled glial fibrillary acidic protein expressing cells were detected 14 days post-SCI, most of which migrated within 70 microm of the region of the central canal. These results show that a minimal SCI adjacent to the ependyma is sufficient to induce an endogenous ependymal cell response where ependymal stem/progenitor cells proliferate and migrate from the region of the central canal, differentiating primarily into astrocytes.

Endogenous and exogenous CNS derived stem/progenitor cell approaches for neurotrauma.

Neural stem/progenitor cells capable of generating new neurons and glia, reside in specific areas of the adult mammalian central nervous system (CNS), including the ependymal region of the spinal cord and the subventricular zone (SVZ), hippocampus, and dentate gyrus of the brain. Much is known about the neurogenic regions in the CNS, and their response to various stimuli including injury, neurotrophins (NFs), morphogens, and environmental factors like learning, stress, and aging. This work has shaped our current views about the CNS's potential to recover lost tissue and function post-traumatically and the therapies to support the intrinsic regenerative capacity of the brain or spinal cord. Recently, intensive research has explored the potential of harvesting, culturing, and transplanting neural stem/progenitors as a therapeutic intervention for spinal cord injury (SCI) and traumatic brain injury (TBI). Another strategy has focused on maximizing the potential of this endogenous population of cells by stimulating their recruitment, proliferation, migration, and differentiation in vivo following traumatic lesions to the CNS. The promise of such experimental treatments has prompted tissue and biomaterial engineers to implant synthetic three-dimensional biodegradable scaffolds seeded with neural stem/progenitors into CNS lesions. Although there is no definitive answer about the ideal cell type for transplantation, strong evidence supports the use of region specific neural stem/progenitors. The technical and logistic considerations for transplanting neural stem/progenitors are extensive and crucial to optimizing and maintaining cell survival both before and after transplantation, as well as for tracking the fate of transplanted cells. These issues have been systematically addressed in many animal models, that has improved our understanding and approach to clinical therapeutic paradigms.

Expression of mRNA encoding extracellular matrix glycoproteins SPARC and SC1 is temporally and spatially regulated in the developing cochlea of the rat inner ear.

SPARC is a multifunctional extracellular matrix (ECM) glycoprotein that shares partial sequence homology with SC1/hevin. These ECM molecules exhibit calcium-binding properties and modulate cellular interactions. This study examines the expression of SC1 and SPARC mRNA in the developing cochlea of the rat inner ear prior to and after the onset of hearing. At all ages examined, SC1 mRNA is highly expressed in neurons of the spiral ganglion. In contrast, SPARC transcripts are not detected in the spiral ganglion but are enriched in the temporal bone and cartilaginous otic capsule surrounding the cochlea. Both SC1 and SPARC mRNA are expressed in connective tissue elements involved in maintaining ionic homeostasis of cochlear fluids. SC1 mRNA is localized to type III fibrocytes of the spiral ligament (slg) and marginal cells of the stria vascularis, while SPARC mRNA is apparent in the spiral limbus and type I fibrocytes of the slg. At postnatal day 10, SPARC mRNA shows a dramatic change in expression. High levels of SPARC transcripts are induced in Deiters cells (dc) of the organ of Corti. Interestingly, this induction of SPARC mRNA correlates with the onset of hearing. This suggests that SPARC may play a role in calcium regulation in dc when functional maturation of the cochlea is attained and rapid changes in calcium levels are required.

Differential mRNA expression of the related extracellular matrix glycoproteins SC1 and SPARC in the rat embryonic nervous system and skeletal structure.

SPARC is a multifunctional extracellular matrix glycoprotein that shares partial sequence homology with SC1. These extracellular matrix molecules are thought to play important roles in modulating cellular interactions. In vitro, SPARC has been shown to exhibit anti-adhesive activity. In the present investigation, in situ hybridization is used to compare the expression patterns of SC1 and SPARC mRNA in the rat embryo. Results show that SC1 and SPARC expression is spatially and temporally regulated. SC1 mRNA is strongly expressed in the embryonic brain and spinal cord, whereas SPARC mRNA is enriched in craniofacial cartilage and skeletal structures. This differential expression pattern in the rat embryo suggests that SC1 plays an important role in the developing nervous system, whereas SPARC participates primarily in events associated with skeletal development. However at embryonic day 17, SC1 and SPARC mRNA show parallel expression patterns in areas of the cerebellum undergoing cell migratory events.

Selective transport of SC1 mRNA, encoding a putative extracellular matrix glycoprotein, during postnatal development of the rat cerebellum and retina.

The selective transport of mRNA species into peripheral processes of cells is an important aspect of gene expression in the nervous system. In this study, we report the transport of SC1 mRNA into the distal processes of Bergmann glial (BG) cells at particular stages of development. SC1 is a putative anti-adhesive extracellular matrix (ECM) glycoprotein that is expressed not only in the developing central nervous system (CNS) but also in the adult brain. The intracellular distribution of SC1 mRNA was examined in two highly laminated neural structures, the cerebellum and retina, during postnatal development and in the adult rat. Our results indicate that SC1 mRNA expression is both spatially and temporally regulated. SC1 message was localized to BG cell bodies at postnatal day 5 (P5) and P10. However, by P15 through to the adult, SC1 mRNA was transported to distal processes of BG cells in the synapse-rich molecular layer (ML) of the cerebellum. In the developing rat retina, SC1 mRNA was expressed in specific neuronal populations by P10, however, transport of SC1 message to the dendrites of these retinal neurons was not detected during development or in the adult. These results indicate neural mechanisms which control the timing and cell type in which selective transport of SC1 mRNA is observed. The localization of SC1 mRNA to the distal processes of BG cells in the synapse-rich ML of the cerebellum could facilitate local control of SC1 protein synthesis which may play roles in synapse formation during development and in synaptic plasticity in the adult.

Phosphinic derivatives as new dual enkephalin-degrading enzyme inhibitors: synthesis, biological properties, and antinociceptive activities.

The development of dual inhibitors of the two zinc metallopeptidases, neprilysin (neutral endopeptidase) and aminopeptidase N involved in the inactivation of the opioid peptides, enkephalins, represents an attractive physiological approach in the search for new analgesics devoid of the major drawbacks of morphine. Phosphinic compounds, corresponding to the general formula H(3)N(+)-CH(R(1))-P(O)(OH)-CH(2)-CH(R(2))-CONH-CH(R(3))-COO(-), able to act as transition-state analogues and to fit the S(1), S(1)', and S(2)' subsites of both enzymes were designed. Selection of the R(1), R(2), and R(3) residues for optimal recognition of these enzymes led to the first dual competitive inhibitors with K(i) values in the nanomolar range for neprilysin and aminopeptidase N. These compounds induce potent analgesic responses after intracerebroventricular or intravenous administrations in mice (hot plate test), and several of them were shown to be, at least, 10 times more potent than the previously described dual inhibitors.

Investigation of subsite preferences in aminopeptidase A (EC 3.4.11.7) led to the design of the first highly potent and selective inhibitors of this enzyme.

The study of the physiological roles of the membrane-bound zinc-aminopeptidase A (glutamyl aminopeptidase, EC 3.4.11.7) needs the design of efficient and selective inhibitors of this enzyme. An acute exploration of aminopeptidase A active site was performed by a combinatorial approach using (3-amino-2-mercapto-acyl)dipeptides able to fit its S(1), S(1)', and S(2)' subsites. This analysis confirmed that the S(1) subsite is optimally blocked by a glutamate or isosteric residues and demonstrated that the S(1)' subsite is hydrophobic whereas the S(2)' subsite recognizes preferentially negatively charged residues derived from aspartic acid. The optimization of these structural parameters led to the synthesis of nanomolar and subnanomolar inhibitors of aminopeptidase A such as H(3)N(+)CH(CH(2)CH(2)SO(3)(-))CH(SH)CO-Ile-(3-COOH)Pro that exhibits a K(i) of 0.87 nM. The best compounds were synthesized by a stereochemically controlled route. These first described highly potent inhibitors could allow studies about the role of physiological substrates of APA such as angiotensin II and cholecystokinin CCK(8) in the central nervous system.