1990 — 1992 |
Fernandez-Valle, Cristina Maria |
F32Activity Code Description: To provide postdoctoral research training to individuals to broaden their scientific background and extend their potential for research in specified health-related areas. |
Expression of Myelin Structural Proteins in Schwann Cell @ University of Miami School of Medicine |
0.901 |
2009 — 2013 |
Fernandez-Valle, Cristina Maria |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Identification of Novel Drug Targets For Use in Preventing Deafness Caused by Nf2 @ University of Central Florida
DESCRIPTION (provided by applicant): Neurofibromatosis type 2 (NF2) is a tumor disorder characterized by development of bilateral vestibular schwannomas also called acoustic neuromas. 98% of all NF2 patients experience partial to complete loss of hearing. Treatment for NF2 is balanced between monitoring tumor growth and the slow but progressive loss of hearing with surgical removal of larger tumors impinging on brainstem function and complete and permanent deafness. Cochlear and auditory brainstem implants have been used to partially restore hearing in a subset of patients with varying success. Therapeutics that slow or reverse tumor growth whilst maintaining hearing are currently lacking. This proposal tests the hypothesis that correcting the cytoskeletal defects in supernumerary schwannoma cells lacking function of the nf2 gene product, schwannomin/merlin, will allow these cells to interact with axons and receive cues promoting their differentiation and/or apoptosis. Identifying proteins that directly bind actin and regulate Schwann cell morphology is of the utmost importance. These proteins can serve as targets for drugs that will repair actin dynamics in schwannoma cells and restore axonal contact. Alternatively, drugs that modify actin regulatory proteins could promote cell death as a result of failed cytokinesis and mitotic spindle organization. One function of schwanomin/merlin is to inhibit Cdc42/Rac activation of p21 activated kinase (PAK). We will investigate LIM kinase (LIMK) and cofilin, terminal targets in a PAK signaling pathway. LIMK is a substrate for PAK, thus its activity is predicted to be high in schwannomas. Cofilin is a ubiquitously expressed actin-binding factor that depolymerizes f-actin and creates nucleation sites for new actin polymerization. Cofilin's function is inhibited by phosphorylation on serine-3 by LIMK. Our preliminary studies demonstrate that LIMK and cofilin modulate actin dynamics and function in Schwann cells. Moreover, our results suggest that LIMK and cofilin act down-stream of Schwannomin/merlin. We propose studies to: 1) identify the role of these proteins in controlling actin polymerization and cellular function in normal rat Schwann cells, 2) establish an in vitro model for NF2 using nf2ex2deleted mouse SCs to determine if inactivation of schwannomin/merlin leads to de-regulation of LIMK and cofilin activity and loss of SC function, and 3) determine if modulators of LIMK and cofilin restore the morphology and function of nf2ex2deleted SCs. These studies are initial steps in validating LIMK and/or cofilin as drug targets for development of an effective treatment for NF2 aimed at preserving hearing. The work in this proposal is relevant to the loss of hearing caused by Neurofibromatosis type 2. This disorder is characterized by development of bilateral acoustic schwannomas and loss of hearing in 98% of patients. As an outcome of this proposal, we hope to create a well characterized in vitro model for NF2 and advance a novel therapeutic direction, the actin modifying proteins LIMK and cofilin.
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0.958 |
2009 — 2010 |
Fernandez-Valle, Cristina Maria |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Mechanisms Modulating Cytoskeletal Dynamics During Schwann Cell Myelination @ University of Central Florida
Myelination allows rapid propagation of action potentials in the nervous system. Dys- and demyelinating disorders are among the most common neurological pathologies affecting young adults. Myelinating glial cells such as Schwann cells of the peripheral nervous system undergo stage specific changes in morphology that enable them to elaborate myelin around an axon. The signaling pathways that regulate actin dynamics in response to extrinsic axonal and extracellular matrix cues are largely unknown. We recently reported that activation of ErbB and B1 integrin receptors induces phosphorylation of Schwannomin, the Neurofibromatosis type 2 tumor suppressor. Schwannomin links receptors to the actin cytoskeleton and modulates the activity of Cdc42/RacGTPase. Thus it is positioned to link receptor activity to actin dynamics and trigger changes in cellular morphology needed for myelination. We hypothesize that Sch modulates actin polymerization through a p21activated kinase (PAK) - LIM kinase - cofilin pathway. In Aim 1, we propose to elucidate the function of cofilin during Schwann cell myelination. We will compare the ability normal and cofilin-deficient Schwann cells to myelinate axons in vitro. In Aim 2, we will assess whether neuregulin and laminin regulate LlMK and cofilin phosphorylation. We will determine whether cofilin is needed for Schwann cells to remodel their plasma membrane in response to NRG and laminin stimulation by conducting live imaging experiments with normal and cofilin-deficient Schwann cells. The immediate outcome of this work will be to identify a novel pathway used by Schwann cells to produce receptor specific changes in morphology associated with myelination. If successful, this work will provide the first complete map of a signaling cascade initiated by axonal and basal lamina ligands that terminates at the final downstream effector protein, actin. This information will impact the identification of increasingly specific drug targets for development of single and combinatorial therapies for myelinating disorders and other Schwann cell disorders such as Neurofibromatosis and Schwannomatosis.
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0.958 |
2010 |
Fernandez-Valle, Cristina Maria |
S10Activity Code Description: To make available to institutions with a high concentration of NIH extramural research awards, research instruments which will be used on a shared basis. |
Zeiss Spinning Disc/Lsm700 Point Scanning Confocal Microscope With Incubation Sys @ University of Central Florida
DESCRIPTION (provided by applicant): This proposal requests funds to purchase a Zeiss Spinning Disc/LSM 700 Point Scanning Confocal Microscope Combination System for the Bioimaging Core Facility of the Burnett School Biomedical Sciences in the newly-formed College of Medicine at University of Central Florida (UCF). The system includes an integrated Zeiss LSM 700 laser scanning confocal microscope, with a cell-observer platform and a CSU-X1 spinning disc system. This instrument allows users to perform live cell time-lapse imaging, 3D imaging, multiple fluorescence and colocalization analyses, nuclear localization, emission fingerprinting, DIC, FRAP, FLIP, photoactivation and photoconversion experiments. The unique capabilities of LSM 700 include variable secondary dichroic beam splitter, FixGate main beam splitter, laser life extender, and integrated software that controls data acquisition and analysis of multiple parameters. There is a critical need for this instrument because there is only one common use confocal microscope available at UCF and in the Central Florida area. It is a Zeiss LSM 510 scanning confocal microscope that was purchased in 2000 and is the sole instrument in the existing Bioimaging Core. This microscope stand has become obsolete and will not be supported by a Zeiss service contract as of September 2009. Furthermore, the current system does not have the capability of live cell imaging or UV excitation of nuclear dyes such as DAPI. The 8 major users of this proposal have 12 NIH funded R01 projects (2 additional R01 will begin in 2009) focused on neurobiology, cardiovascular diseases, infectious diseases, and cancer. Each award has experiments that require confocal and live imaging. In addition, there are 4 minor users who have NIH-funded projects. The purchase of the microscope system will help sustain existing NIH-funded research programs and will help recruit additional outstanding biomedical researchers. This will help the unit fulfill its strategic goal defined in 2003 to recruit 34 new faculty to build a well-funded nationally recognized biomedical research program at UCF.
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0.958 |
2018 |
Fernandez-Valle, Cristina Maria |
R56Activity Code Description: To provide limited interim research support based on the merit of a pending R01 application while applicant gathers additional data to revise a new or competing renewal application. This grant will underwrite highly meritorious applications that if given the opportunity to revise their application could meet IC recommended standards and would be missed opportunities if not funded. Interim funded ends when the applicant succeeds in obtaining an R01 or other competing award built on the R56 grant. These awards are not renewable. |
Co-Targeting Fak and Pi3k For Nf2 Schwannomas @ University of Central Florida
Mutations in the merlin tumor suppressor gene (NF2) cause the rare tumor disorder Neurofibromatosis Type 2 (NF2). Individuals with NF2 develop schwannomas as well as meningiomas and ependymomas throughout their lifetime. Bilateral vestibular schwannomas (VS) alone are diagnostic for NF2 and are typically not removed until hearing worsens or there is risk of brainstem compression. Resection of VS however leaves patients deaf and often with facial paralysis and balance deficits. Thus there is a great need for pharmaceuticals to slow or prevent schwannoma growth that are safe for chronic use. We adopted an unbiased chemical genomics approach to identify drug targets and compounds/drugs for NF2 schwannoma therapy. For this work we created merlin- deficient mouse and human Schwann cell lines, optimized their use in high-throughput and high-content assays, and screened commercial and custom compound libraries using robotic platforms. The results identified several PI3K and PI3K/mTOR inhibitors that selectively reduce viability of merlin-deficient mouse Schwann cells compared to wild-type Schwann cells. Kinome analysis of merlin-deficient human Schwann cells exposed to PI3K inhibitors for 24 hours revealed that cells consistently increased activity of focal adhesion kinase (FAK) and SRC kinase family members. Rewiring of kinase networks occurs in cancer cells developing drug resistance to monotherapies. Moreover, a drug combination screen revealed maximal synergy between a PI3K inhibitor and a FAK inhibitor in selectively reducing viability of a human merlin-deficient Schwann cell line. These results suggest that targeting the FAK/SRC pathway in combination with PI3K inhibitors should provide sustained inhibition of merlin-deficient Schwann cell proliferation and/or survival. In this proposal, we advance these findings by investigating the efficacy of combined PI3K pathway and FAK/SRC inhibition in cell based assays using mouse and human merlin-deficient Schwann cell lines, as well as patient-derived VS cells, and in vivo using allograft and genetic models of NF2 schwannomas. The aims of the proposal are to: 1) screen combinations of PI3K pathway and FAK/SRC inhibitors for synergistic efficacy in reducing viability of merlin- deficient Schwann cell lines; 2) pre-screen advanced drug combinations for in vivo efficacy using a rapid sciatic nerve allograft model, 3) test efficacy of one drug combination in preserving hearing and balance, and slowing schwannoma growth in a genetically engineered mouse model, and 4) assess successful drug combinations in primary human VS cells. We expect to obtain the necessary pre-clinical data to support the potential use of PI3K pathway and FAK/SRC inhibitors, alone or in combination for slowing progression of hearing loss and schwannoma growth in NF2. Moreover these studies will establish a streamlined in vitro and in vivo drug testing platform for future studies.
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0.958 |
2018 — 2021 |
Fernandez-Valle, Cristina Maria Liu, Xue Z (co-PI) [⬀] |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Preclinical Testing of Pi3k Inhibitors For Vestibular Schwannomas @ University of Central Florida
ABSTRACT Mutations in the merlin (NF2) tumor suppressor gene cause the benign tumor disorder, Neurofibromatosis type 2 (NF2). This disorder predisposes individuals to develop bilateral vestibular schwannomas (VS) that cause progressive hearing loss and can cause life-threatening brainstem compression. Because surgical removal of a VS often causes deafness, facial paralysis, and imbalance, there is a need to develop drug therapies to slow or prevent VS growth and preserve nerve function. We have worked to establish an in vitro and in vivo drug screening platform to identify novel compounds as well as FDA-approved drugs that can be developed/repurposed for VS therapies. Toward this goal, we have created mouse and human merlin-deficient Schwann cell lines and optimized their use in 384-well high-throughput and high-content assays in order to screen large compound/drug libraries using robotic platforms. This approach identified several phosphoinositide- 3 kinase (PI3K) inhibitors that selectively reduce viability of mouse merlin-deficient compared to wild-type Schwann cells with nanomolar IG50. This initial finding was confirmed in human merlin-deficient Schwann cell lines for multiple PI3K, dual PI3K/mTOR and PI3K/HDAC inhibitors. Because PI3K plays a critical role in cell proliferation, survival, and invasion, there are currently 15 different PI3K inhibitors in clinical trials for various blood cancers and solid tumors. The first in class PI3K inhibitor (idelalisib) was approved in 2014 for leukemia. In this proposal, we advance our findings by conducting a systematic screen of PI3K pathway inhibitors in vitro and in vivo. The aims of this proposal are to: 1) profile a library of PI3K pathway inhibitors for efficacy in reducing viability of human merlin-deficient Schwann cell lines and primary human VS cells; 2) test efficacy of the advanced PI3K inhibitors to slow graft expansion and preserve hearing and balance in a novel rat xenograft model, and 3) conduct phenotypic, kinome, and transcriptome analysis to reveal the molecular signatures and adaptive changes of the cells and grafts to the inhibitors. We expect to obtain the necessary pre-clinical data to support the potential use of PI3K inhibitors in patients with NF2-associated VS.
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0.958 |