Montana Molecular is in the business of developing and marketing genetically encoded biosensors for academic research and drug discovery in living cells. Our biosensors provide robust detection of crucial second messenger signaling, including DAG, PIP2, cAMP and Ca2+ on both fluorescence microscopes and the large installed base of automated plate readers, with metrics that match or surpass those of traditional biochemical and luminescence based assays. We have an established customer base, with over 100 early adopters in academia and industry. Our customers and industry partners have indicated a growing need for assays capable of targeting specific populations of living neurons and for ?assay-ready? iPSC derived neuronal lines for use in academia and secondary drug screens. Genetically encoded assays for GPCR signaling, developed by Montana Molecular, fit seamlessly with these needs. They can be used for kinetic measurements in living cells, can be targeted to specific populations of cells in complex tissue, and have been independently validated on a wide variety of fluorescence microscopes and automated plate readers. To meet the needs of our customers and to penetrate the growing human iPSC market, we propose two specific aims.
In Aim 1 we will engineer a new baculovirus vector that circumvents the payload limits and safety considerations of Lentivirus and AAV.
In Aim 2, we will partner with iPSC developers in both academia and industry to create the first assay-ready iPSC derived neurons stably expressing biosensors for the detection of multiple facets of GPCR signaling.

Public Health Relevance

Investigations of cell signaling and drug discovery have largely been limited to highly engineered cell lines, that are ultimately quite different from the human condition. The goal of this PhaseIIB proposal is to develop technologies that will allow neuroscientists and drug discovery teams to perform their studies, with an unprecedented level of precision, in cells and tissues that are directly relevant to human disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Small Business Innovation Research Grants (SBIR) - Phase II (R44)
Project #
2R44NS082222-04
Application #
9346434
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Fertig, Stephanie
Project Start
2014-01-15
Project End
2019-08-31
Budget Start
2017-09-15
Budget End
2018-08-31
Support Year
4
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Montana Molecular, LLC
Department
Type
DUNS #
145280157
City
Bozeman
State
MT
Country
United States
Zip Code
59715
Moore, Bryn S; Stepanchick, Ann N; Tewson, Paul H et al. (2016) Cilia have high cAMP levels that are inhibited by Sonic Hedgehog-regulated calcium dynamics. Proc Natl Acad Sci U S A 113:13069-13074
Tewson, Paul H; Martinka, Scott; Shaner, Nathan C et al. (2016) New DAG and cAMP Sensors Optimized for Live-Cell Assays in Automated Laboratories. J Biomol Screen 21:298-305
Ding, Yidan; Li, Jiao; Enterina, Jhon Ralph et al. (2015) Ratiometric biosensors based on dimerization-dependent fluorescent protein exchange. Nat Methods 12:195-8
Chen, Xingjuan; Egly, Christian; Riley, Ashley M et al. (2014) PKC-dependent Phosphorylation of the H1 Histamine Receptor Modulates TRPC6 Activity. Cells 3:247-57
Tewson, Paul H; Quinn, Anne Marie; Hughes, Thomas E (2013) A multiplexed fluorescent assay for independent second-messenger systems: decoding GPCR activation in living cells. J Biomol Screen 18:797-806
Tewson, Paul; Westenberg, Mara; Zhao, Yongxin et al. (2012) Simultaneous detection of Ca2+ and diacylglycerol signaling in living cells. PLoS One 7:e42791