The chemical transformations of biology occur in a compartmentalized context inside the cell membrane. Systematically studying this compartmentalized chemistry and harnessing its benefits for therapeutic applications through directed enzyme evolution will require methods for controlled synthesis and functional screening of cell-like compartments. Mentored research activities significantly expanded on current efforts in microfluidic directed evolution by exploring circuitry for the controlled high-throughput synthesis of monodisperse water droplets in oil for in vitro compartmentalization (IVC). This strategy Is enabling new explorations of RNA's catalytic fitness landscape by prohibiting a single advantageous genotype from dominating in the selective amplification reaction, and exaggerating neutral drift ofthe population. A nozzle array microfluidic IVC (MIVC) circuit was developed for these experiments and enabled selections encompassing l e 8 individuals per hour. Directed evolution of proteins with complex phenotypes (transport, membrane display, catalysis) will form the theme for independent phase investigations. The pIVC system will be used to synthesize monodisperse lipid vesicles for compartmentalization and functional display of integral membrane proteins, P-galactosidase and hemolysin will serve as models for using the pIVC processor to evolve new catalytic and selective transport functions on cytosolic and transmembrane proteins, respectively. Long-tennn research program goals include evolving membrane receptors (CCR5 and CD4) in lipid vesicles, selecting for enhanced binding of viral protein-receptor complexes, evolutionary structure-function studies, and synthesizing membrane-bound evolvable ligands for applications In targeted and decoy therapeutics.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Specialized Center (P50)
Project #
5P50CA083636-15
Application #
8523022
Study Section
Special Emphasis Panel (ZCA1-RPRB-M)
Project Start
Project End
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
15
Fiscal Year
2013
Total Cost
$272,427
Indirect Cost
$81,314
Name
Fred Hutchinson Cancer Research Center
Department
Type
DUNS #
078200995
City
Seattle
State
WA
Country
United States
Zip Code
98109
Wang, Yifan; Krais, John J; Bernhardy, Andrea J et al. (2016) RING domain-deficient BRCA1 promotes PARP inhibitor and platinum resistance. J Clin Invest 126:3145-57
Norquist, Barbara M; Harrell, Maria I; Brady, Mark F et al. (2016) Inherited Mutations in Women With Ovarian Carcinoma. JAMA Oncol 2:482-90
Yu-Rice, Yi; Edassery, Seby L; Urban, Nicole et al. (2016) Selenium Binding Protein 1 autoantibodies in ovarian disorders and ovarian cancer. Reproduction :
Yumul, Roma; Richter, Maximilian; Lu, Zhuo-Zhuang et al. (2016) Epithelial Junction Opener Improves Oncolytic Adenovirus Therapy in Mouse Tumor Models. Hum Gene Ther 27:325-37
Wang, Yifan; Bernhardy, Andrea J; Cruz, Cristina et al. (2016) The BRCA1-Δ11q Alternative Splice Isoform Bypasses Germline Mutations and Promotes Therapeutic Resistance to PARP Inhibition and Cisplatin. Cancer Res 76:2778-90
Andersen, M Robyn; Thorpe, Jason; Buist, Diana S M et al. (2016) Cancer Risk Awareness and Concern among Women with a Family History of Breast or Ovarian Cancer. Behav Med 42:18-28
Buas, Matthew F; Gu, Haiwei; Djukovic, Danijel et al. (2016) Identification of novel candidate plasma metabolite biomarkers for distinguishing serous ovarian carcinoma and benign serous ovarian tumors. Gynecol Oncol 140:138-44
Elias, Kevin M; Emori, Megan M; Westerling, Thomas et al. (2016) Epigenetic remodeling regulates transcriptional changes between ovarian cancer and benign precursors. JCI Insight 1:
Gregory, Mark T; Bertout, Jessica A; Ericson, Nolan G et al. (2016) Targeted single molecule mutation detection with massively parallel sequencing. Nucleic Acids Res 44:e22
Bernards, Sarah S; Norquist, Barbara M; Harrell, Maria I et al. (2016) Genetic characterization of early onset ovarian carcinoma. Gynecol Oncol 140:221-5

Showing the most recent 10 out of 167 publications