This is a renewal request to continue work to identify the sites on tubulin which interact with the antitumor drug paclitaxel through the use of photo affinity and electrophilic analogs synthesized in the Co-Principal Investigator s laboratory and to study tubulin- paclitaxel interactions using fluorescent analogs also prepared in the Co-PI s laboratory. Analogs with reactive groups at different positions around the paclitaxel molecule are expected to allow the researchers to identify peptides in the binding domain around the taxane molecule. Azido derivatives of the 2-and 3'-N-benzoyl groups have been prepared and found to have good biological activity. These derivatives have been synthesized in radio-labeled form and are being used in photo labeling studies. It is proposed to synthesize additional reactive derivatives at the 7,-10-and 3'-positions and use those to label tubulin. After labeling, peptides are produced by chemical and proteolytic procedures and purified by a combination of chromatographies. Sequences and sites of modification will be determined by gas phase sequencing and mass spectrometry. As a compliment to the photo-induced labeling experiments, several derivatives containing electrophilic substituents on paclitaxel will be utilized to label the binding domain on tubulin. Finally, newly prepared fluorescent analogs will be utilized to evaluate the polarity of the binding domain and the extent bound paclitaxel is exposed to the solvent matrix. A greater understanding of paclitaxel-tubulin interactions should be helpful for the development of new derivatives with better activity and less toxicity.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA055141-07
Application #
2894895
Study Section
Special Emphasis Panel (ZRG3-BNP (02))
Program Officer
Lees, Robert
Project Start
1993-01-01
Project End
2001-04-30
Budget Start
1999-05-01
Budget End
2001-04-30
Support Year
7
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Kansas Lawrence
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
072933393
City
Lawrence
State
KS
Country
United States
Zip Code
66045
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Gupta Jr, Mohan L; Bode, Claudia J; Thrower, Douglas A et al. (2002) beta-Tubulin C354 mutations that severely decrease microtubule dynamics do not prevent nuclear migration in yeast. Mol Biol Cell 13:2919-32
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Gupta, M L; Toso, R J; Farrell, K W et al. (1995) Commercial [3H]glutamate contains a contaminant that labels tubulin covalently. Anal Biochem 230:350-3
Sengupta, S; Boge, T C; Georg, G I et al. (1995) Interaction of a fluorescent paclitaxel analogue with tubulin. Biochemistry 34:11889-94

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