Three-Dimensional Visualization and Volumetric Analysis of Cancer Therapeutic Agents and Cell Interactions Using Raman Imaging, 10-9110

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Principal Investigator
Jian Ling

Inclusive Dates: 01/01/99 - 06/30/00

Background - The determination of drug mechanisms at the cellular level is a problem of great importance to the evaluation and development of new anticancer agents. Recently, the implementation of rational drug design, combinatorial chemistry techniques, and high-throughput screening has led to large numbers of new potential drugs. Currently there are limited cost-effective and efficient approaches to understanding how these potential drugs work at the cellular level. This lack of understanding requires pharmaceutical companies to spend millions of dollars in animal and clinical studies to assess a candidate drug

Approach - . The objective of this project was to develop direct Raman imaging, a cost-effective tool, for the study of drug mechanisms at the cellular level. The application of direct Raman imaging was studied to determine drug distribution in single living cells. To extract useful information from a noisy Raman image, a model was developed to describe the degradation of Raman signals by several processes: nonuniform illumination of the laser-excitation source, distortion by the microscope system, and the influence of additive signal-dependent Gaussian noise that arises during direct Raman imaging. Using this model, special-purpose image-processing algorithms were developed to restore the Raman images. Taxol, an important anticancer agent whose mechanisms at the cellular level have been well studied, was used to evaluate the capabilities of direct Raman imaging. Raman images were obtained from an MDA-435 cancer cell before and after the drug treatment. The ratio image between the two images indicates the two-dimensional distribution of taxol within the cell.

Accomplishments - From the Raman images, taxol was found to have high intracellular concentration near the plasma membrane of MDA-435 breast tumor cells, as indicated in the illustrations below. This finding is well explained by the binding characteristics of the taxol and its molecular target - the microtubules. This result demonstrated the feasibility of using direct Raman imaging to determine the intracellular distribution of a drug. From the drug distribution, its mechanism can be studied.

(a)

(b)

 
(a) White light image of a breast tumor cell before exposure to taxol, an anticancer agent; (b) ratio Raman image [corresponding to the red-box area in (a)] recorded one hour after exposure to taxol. Most of the taxol was concentrated near the cell membrane. The color bar indicates the relative intensity of the Raman signal (the highest intensity is at the top of the bar).

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