Tubulin Inhibitors: Synthesis of Combretastatin A-4 Analogs

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Tubulin Inhibitors Synthesis of Combretastatin
A-4 Analogs
Structure of CA4

• By Lauren Lee Jenna Vanderham
• Dr. Moses Lee
• Hope College

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Tubulin and The Premier Tubulin
InhibitorCombretastatin

• Tubulin is an essential component of the
  cytoskeleton in cells.
• CA4 is a natural product, which is extracted from
  an African Willow Plant
• CA4-Clinical anti-tumor agent
• Inhibits tubulin polymerization
• CA4 has Solubility problems
• To increase solubility, a Phosphate was added to
  the OH group of CA4 to create a pro-drug. This
  pro-drug is now in phase II clinical trails
• CA4 can be modified

African Willow Plant (Combrettum Caffrum)
Combretastatin A4 (CA4)
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Previous Work


Pyrazoline
twisted
CA-4
Pyrazole
twisted
flat
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Premier Compound


LL-A-99
IC500.52µM (L1210)
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Project I- Thiourea analogs


R
R
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Project I- Thiourea analogs cont.


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Making C-A4 Analogs


Thiourea
2.5 eq. Thiourea
2.5 eq. K2CO3 Reflux in ethanol overnight
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Cytotoxicity Testing

• Cell lines L1210 B16
• Use MTT assay
• MTT solution is yellow
• MTT reduced in viable cells
• Product formazan is purple

Increasing Drug
IC50 Drug concentration required to reduce cell
population by 50 relative to untreated control
Spectrophotometer

• MTT 3-(4,5-dimethylthiazol-2-yl)diphenyltetrazoli
  um bromide

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Results of Current Work


IC50 (µM) (L1210)
0.52
32
48.3
gt100
5.5
56.3
gt100
gt100
gt100
IC50 (µM) (L1210)
0.426
2.146
strongly inhibited tubulin polymerization in rat
aorta cells.
The IC50 value of CA4 is 0.007 µM
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Project II- DNA Binders
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Project II- DNA Binders
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Previous Work
Furamidine Minor Groove AT rich sequences Wilson
et al. GSU Atlanta
Minor Groove AT rich sequences Lee at al. Furman
University, Greenville SC
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Procedure
EtOH NaOH

NH2NH2 Acetic Acid Reflux 3 hrs.
O
1. CH3C-Cl, MeOH O?C - r.t. 2. MeOH, NH3 -78?C -
r.t.

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Nitrogen Mustard

Covalent bond to DNA
Positive Charge
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Future Work

• Finish Pinner Reaction for LL-A-133, the nitrogen
  mustard
• Jen Ruprich
• Test the DNA binding of the molecules
• Alan Sielaff
• Get the Mass Spectrometry Data from the
  University of South Carolina

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Acknowledgments

• Dr. Moses Lee/ Dr. Toni Brown/Dr. Hilary Mackey
• Lee Lab Mates
• Ryan Davis, Jen Ruprich, Alan Sielaff
• Dr. Susan Mooberry, Southwest Foundation for
  Biomedical Research
• Hope College

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