Recommended
More Related Content
Similar to 2015 SAHA presentation
Similar to 2015 SAHA presentation (20)
2015 SAHA presentation
- 1. VORINOSTAT INDUCES P-GLYCOPROTEIN ACTIVITY IN BLOOD-BRAIN BARRIER ENDOTHELIUM Taylor Liles
- 2. OUTLINE • 1. Introduction • a. Blood brain barrier: Structure and Function • b. ABC Efflux transporters and their role in BBB function • c. Vornistat • 2. Hypothesis • 3. Experiments and Data • 4. Conclusion and Outlook • 5. Acknowledgements
- 3. THE BLOOD BRAIN BARRIER • Physical and chemical barrier • 3D culture • Tight Junctions • Efflux Transporters https://loonylabs.files.wordpress.com/2015/03/blood-brain-barrier.jpg?w=590 https://www.google.com/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAcQjRxqFQoTCJLS25zV58YCFQY6kgod1ycM0w&url=http%3A%2F%2 Fwww.pubfacts.com%2Ffulltext_frame.php%3FPMID%3D23678437%26title%3DImpacts%2520of%2520blood- brain%2520barrier%2520in%2520drug%2520delivery%2520and%2520targeting%2520of%2520brain%2520tumors.&ei=JNerVZKwN4b0yATXz7CYDQ&bvm=bv.98197061, d.cGU&psig=AFQjCNHtqILV8_qvW9ud2gAHlLbiUjz-Yw&ust=1437411150229860
- 4. P-GP (MDR1) • ATP-binding cassette Transporter • Broad range of substrate • Lipids • Xenobiotics • Peptides • Uncharged or positive charge • Hydrophobic • Intestines, liver, kidneys, and brain micro- vasular endothelium https://upload.wikimedia.org/wikipedia/commons/thumb/f/fe/MDR3_3 g5u.png/500px-MDR3_3g5u.png
- 5. VORNISTAT (SAHA) • Non-Specific Histone Deacetylase (HDAC) Inhibitor • FDA approval in 2006 for persistent or recurrent Cutaneous T-Cell Lymphoma
- 6. VORNISTAT (SAHA) • Causes death and differentiation in cancer cells ¹ • Cellular changes through non-histone proteins ² • Increase synaptic activity ³ • Low brain penetration ³ • P-GP substrate ³ 1. Marks PA. Discovery and development of SAHA as an anticancer agent. Oncogene. 2007 Feb 26;26(9):1351-6. Review. PubMed PMID: 17322921. 2. Spange S, Wagner T, Heinzel T, Krämer OH. Acetylation of non-histone proteins modulates cellular signalling at multiple levels. Int J Biochem Cell Biol. 2009 Jan;41(1):185-98. doi: 10.1016/j.biocel.2008.08.027. Epub 2008 Sep 2. Review. PubMed PMID: 18804549. 3. Hanson, J. E., La, H., Plise, E., Chen, Y.-H., Ding, X., Hanania, T., … Zhou, Q. (2013). SAHA Enhances Synaptic Function and Plasticity In Vitro but Has Limited Brain Availability In Vivo and Does Not Impact Cognition. PLoS ONE, 8(7), e69964. doi:10.1371/journal.pone.0069964
- 7. HYPOTHESIS • Can SAHA modulate activity and/or expression of P-gp transporters in Brain microvascular endothelial cells?
- 8. METHODS • Mouse Primary and Human D3 cell line BMECs • Increasing concentrations of SAHA over 1-6 hour time points • ATP-ase assay • P-GP GLO assay • Rhodamine efflux activity assay 1 hour Crtl SAHA 10 nm SAHA 100 nm SAHA 1000 nm 3 hours Crtl SAHA 10 nm SAHA 100 nm SAHA 1000 nm 6 Hours Crtl SAHA 10 nm SAHA 100 nm SAHA 1000 nm
- 9. VIABILITY ASSAYS: D3 CELLS ATP ASSAYLDH ASSAY
- 10. PGP ATP-ASE ASSAY • Assesses consumption of ATP • Metabolic activity
- 11. RHODAMINE ASSAYS • Rhodamine-123 is a lipid-soluble, fluorescent dye • Cells undergo experimental conditions and then are incubated in a R123 solution • The solution is removed, the cells washed, and a fluorescent reading is taken
- 12. RHODAMINE ASSAYS: D3 CELLS
- 13. RHODAMINE ASSAYS: MOUSE PRIMARY CELLS
- 14. CONCLUSION •Vornistat does seem to exert an enhancing effect on P-gp activity. •This could disqualify Vornistat as a potential therapeutic in Brain Cancers. •Further tests need to be performed assessing other efflux transporters.
- 15. ACKNOWLEDGEMENTS • Ravi Sajja Research Scientist • Shikha Prasad Graduate Student • Mohammad Kaisar Graduate Student • Luca Cucullo Assistant Professor •Teressa Carlisle GPPS, GSBS, ABRI Coordinator All involved in organizing, funding, and speaking for the ABRI Program Thank you for investing in my success as a scientist!
- 16. QUESTIONS?