The goal of this proposal is to develop microfluidic devices for effective single-cell analysis in biomedical laboratories without the requirement of specialized expertise. The lack of ability to perform biochemical and molecular characterization at the single-cell level is a critical obstacle in cancer research and diagnosis. We have developed microfluidic devices for efficient large-scale single-cell analysis to overcome this obstacle. The technology has broad applications in cancer research and diagnosis for identification, enumeration and characterization of the rare cells including 1) circulating tumor cells (CTC) in occult metastases;2)T-regulatory cells, 3) tumor-specific T-cytotoxic cells;4) cancer stem cells;5) malignant cells captured by laser capture microdissection (LCM);and 6) suspicious malignant cells in needle biopsy. Isolation methods have been developed to capture rare cancer cells from blood and other tissues for cancer diagnosis. However, there is still a lack of robust methods to perform biochemical and molecular characterization on these captured cells at the single-cell levels. Here, we propose to develop an inexpensive and integrated microfluidic device that can simultaneously profile 100 single-cells within 3 hours. The proposed device can be fabricated and used in general laboratories without the need of specialized expertise, and has a 5 fold higher mRNA-to-cDNA efficiency than current bulk assays. With our microfluidic devices, multiple biochemical analyses can now be performed at the single-cell levels with a limited number of target cells such as the rare cancer cells listed above. We propose to optimize our devices for single-cell biochemical characterization of CTC captured by our membrane filters. Characterization of these captured CTC is likely to lead to the identification of molecular therapeutic targets. After developing and optimizing the device with this R21 proposal, a R33 proposal will be submitted to apply this device for molecular characterization of CTC captured from bloods of cancer patients at the single-cell level.

Public Health Relevance

We proposed to develop an inexpensive integrated microfluidic device for large scale single-cell gene expression profiling. This device will permit analysis of hundreds to thousands rare cancer cells at the single-cell level. Once developed, this device will be a novel platform for molecular characterization of rare cancer cells such as circulating tumor cells (CTC) captured from patient bloods.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21CA134391-01A1
Application #
7630366
Study Section
Special Emphasis Panel (ZCA1-SRRB-4 (J1))
Program Officer
Rasooly, Avraham
Project Start
2009-06-01
Project End
2011-05-31
Budget Start
2009-06-01
Budget End
2010-05-31
Support Year
1
Fiscal Year
2009
Total Cost
$212,715
Indirect Cost
Name
University of Southern California
Department
Pathology
Type
Schools of Medicine
DUNS #
072933393
City
Los Angeles
State
CA
Country
United States
Zip Code
90089
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Vu, Long T; Keschrumrus, Vic; Zhang, Xi et al. (2015) Tissue elasticity regulated tumor gene expression: implication for diagnostic biomarkers of primitive neuroectodermal tumor. PLoS One 10:e0120336
Zhang, Xi; Zhang, Cheng; Li, Zhongjun et al. (2013) Investigating evolutionary perspective of carcinogenesis with single-cell transcriptome analysis. Chin J Cancer 32:636-9
Chen, Yan; Zhang, Baoyue; Feng, Hongtao et al. (2012) An automated microfluidic device for assessment of mammalian cell genetic stability. Lab Chip 12:3930-5
Fan, Jian-Bing; Chen, Jing; April, Craig S et al. (2012) Highly parallel genome-wide expression analysis of single mammalian cells. PLoS One 7:e30794
Li, Shengwen Calvin; Jin, Yangsun; Loudon, William G et al. (2011) Increase developmental plasticity of human keratinocytes with gene suppression. Proc Natl Acad Sci U S A 108:12793-8
Xu, Xiaowei; Zhong, Jiang F (2010) Circulating tumor cells and melanoma progression. J Invest Dermatol 130:2349-51
Li, Shengwen Calvin; Zhong, Jiang F (2009) Twisting immune responses for allogeneic stem cell therapy. World J Stem Cells 1:30-35