IS PLATINUM-BASED DRUG CYTOTOXICITY ENHANCED BY

THE ANTIDEPRESSANT DESIPRAMINE?

Sandeep SharmaVCU/HHMI Summer Scholars program

Mentor: Dr J.J. RyanDepartment of Biology

Hypothesis• Since resistance to chemotherapeutic drugs

is frequently associated with reduced drug uptake, increasing drug uptake can enhance cell killing. Some anti-depressants can alter the function of transport proteins that allow cells to accumulate drugs. We postulated that the anti-depressant desipramine could enhance cell killing by platinum-based chemotherapy.

• The two major experiments carried out were:

• The Effect of Desipramine on cisplatin (c DDP)-mediated killing of A2780 ovarian carcinoma cells.

• Effect of Desipramine on Oxaliplatin + 5FU- Mediated killing of HCT116 colorectal carcinoma cells.

Cis-diamminedichloroplatinum(II) c DDP

• A platinum based chemotherapy drug used to treat several types of cancer such as sarcomas, lyphomas and ovarian cancer.

• It was the first member of its class which also includes carboplatin and oxaliplatin.

• The cytoxoxicity is mainly through the interactions with DNA and the inhibtion of synthesis and replication by formation of interstrand and

intrastrand cross links.

Background

Oxaliplatin

• Is a platinum based chemotherapy drug. Typically is administered with fluorouracil and leucovorin for the treatment of colorectal cancer.

• It has a similar structure to cisplatin where the two amine groups are replaced by cyclohexyl diamine and the chlorine ligands are replaced by the oxalato-bidentate.

Fluorouracil 5-FU

• Belongs to the family of drugs called antimetabolites. It acts prinicipally as a thymidylate synthase inhibitor, interrupting an enzyme that is critical factor in the synthesis of pyrimidine which is important in DNA replication.

• Desipramine- A tricyclic antidepressant. Its activity rises from inhibiting reuptake and in the process raising the level of seretonin and norepinephrin in the synaptic cleft.

• It is also used for treating neuropathic pain especially with prostate cancer patients.

• Desipramine can block organic ion transporters (OCTs), and hence may alter drug uptake.

Desipramine

PROTOCOL

• Tryposonized and collected HCT 116 cells and A2780 cells.

• Aliqouted 150,00 cells into 3mL cRPMI(media) in 6 well plates.

• Added the different drugs according to the different culture conditions.

• Cultured the cells for 72 hours.• Collected adherent and non-adherent cells.• Fixed the cells in ethanol.• Performed PI-DNA staining.• Analysed the cells for DNA content on a

FACScan/FACSCalibur flow cytometer.

PI DNA STAINING ASSAY

Treatment with Drug

CellsFixed

overnight

Stained with propidium

iodide

Analyzed by Flow Cytometry

Cell Cycle

0 1 2 3 4 5 6

0

25

50

75

100

cDDP

cDDP + Desipramine

Desipiramine

Cisplatin (M)

Desipramine Augments c DDP Induced Killing

Control Oxali+5FU Desipramine Oxali+5FU + Desip0

5

10

15

20

25

Cell Killing with Desipramine is Better than Oxaliplatin and 5FU Alone

Conclusions

• Desipramine augments killing of both ovarian and colorectal carcinoma cells.

• Other studies from our lab recently showed that desipramine has little or no effect on drug uptake, but enhances the activation of apoptotic pathways, including p53.

• Because desipramine is approved for clinical use, it may be an excellent means to augment standard chemotherapeutic treatment.

Acknowledgements

• Dr. Ryan for his tremendous support and guidance.• All members of the Ryan lab for all their help and

encouragement:• Peyman Kabolizadeh• Brian Barnstein• Sarah Kennedy• Jenifer Brenzovich• Kevin Dholaria

• Dr. Buck and Dr. Johnson.