Date of Award

Spring 1985

Project Type

Dissertation

Program or Major

Biochemistry

Degree Name

Doctor of Philosophy

Abstract

The induction of mammary cancer in female Sprague-Dawley rats by a single of 1-methyl-1-nitrosourea was used as an experimental model system for identifying an effective agent in the prevention of breast cancer. The polyamine concentrations in whole mammary gland or mammary epithelial cells remained unchanged in the post-initiation stages of mammary carcinogenesis prior to the appearance of detectable tumors. However, mammary tumors had much higher concentrations of polyamines as compared to normal mammary gland. The development of chemically-induced carcinomas in rats was dramatically suppressed by the provision of a 1% solution of D,L-(alpha)-difluoromethyl-ornithine (DFMO), an inhibitor of polyamine biosynthesis, in drinking water for 6 months. This treatment significantly reduced cancer incidence and the average number of cancers per rat and prolonged cancer-free time. The average weight of mammary gland tumors induced in animals receiving DFMO was reduced ten-fold in comparison to tumors occurring in untreated rats. The concentrations of polyamines and ornithine decarboxylase activity in the tumors of rats receiving 0.5% DFMO were significantly reduced in comparison with data obtained in tumors of control animals. DFMO did not affect polyamine concentrations in the mammary gland of saline-treated animals. DFMO appeared to block some aspect of promotional stage(s) of mammary carcinogenesis dependent upon polyamines.

To study the molecular mechanism by which DFMO acts as an antiproliferative agent in carcinogenesis, HeLa cells were used. HeLa cells were synchronized by the double thymidine block procedure and depleted of polyamines by the use of 1 mM DFMO. These polyamine-depleted cells exhibited a severe deficiency in S phase DNA synthesis and cell proliferation. This inhibitory effect of DFMO could be partially reversed by the addition of polyamines to the cell cultures at the beginning of S phase. Polyamine-deficient cells did not lack any essential cytosol factor(s) for DNA synthesis. Furthermore, salt extracts of nuclei containing DNA polymerase activity did not restore DNA synthesis in nuclei of polyamine-deficient cells. Polyamines may be required for the synthesis of molecules or for their interaction in control mechanisms preceding S phase DNA synthesis.

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