Tobacco Documents Online

THE COUNCIL FOR TOBACCO RESEARCH-U.S.A.. INC. 900 THIRD AVENUE NEW YORK. N.Y. 10022 Memorandum To: Dr. J. F. Glenn and Staff From: D.H.Ford Re: Site visit with Dr. Y. Osawa, Medical Foundation of Buffalo, N.Y., October 6, 1988 Grant No. 1643B entitled "Aromatase inhibitors in cigarette smoke and tobacco." Goals: Since the serendipitous discovery of the presence of a substance in cigarette smoke and in raw tobacco which inhibits aromatase activity, Dr. Osawa and colleagues have focused their attention to determining the nature of this compound and on whether or not it could be an important pharmaceutical agent in preventing estrogen dependent cancers. Results: An intensive series of biochemical studies has revealed that the substance in smoke which inhibits aromatase is an N-acylnornicotine. He then synthesized a number of derivitives o-f nornicotine, obtaining 5 which demonstrated a high level of inhib- ition. The two most active, N-(4-hydroxy-n-undecanoyl) and N-n-decanoylnornicotine were found to be toxic in both in vitro and in vivo studies, while the N-n-octanoylnornicotine, with a much lower potency was still a highly effective inhib"itor of aromatase. Injection of this derivitive into BUF/N female rats with regular 4 day estrus cycles lengthened the time between ovulatory periods for 3 cycles. This was accomplished by a single injection of 50mg/kg on the day of proestrus. It also showed a significant effect in prevent- ing _ breast cancer in this strain of rats following treatment with NMU. In conjunction with Dr. Callard, Dr. Osawa has observed the presence of an aromatase isozyme in the brain of goldfish synapto- somes. This enzyme was shown to be an effective aromatase in an in vitro mammalian system and its activity was suppressed by his anti-aromatase. It is of interest to note that this aromatase was present in significant amounts only during the spawning period. Since brains from both male and female fish were considered together, it is not known if there is a sex difference. Studied with adult rat brains indicated that there were only very low levels present. However, since the major role for aromatase in the male rat brain occurs only in the first 5 to 7 days after birth when its conversion of testosterone to estrogen is necessary for masculinization of the male brain, it is quite possible that significant levels of this enzyme might be present only during this period. Further, since the CNS region in which this hormone is required at this time is restricted to the preoptic region of the hypothalamus, the analysis of whole adult brains might well have diluted any concentration of aromatase which might have been present. Dr. Osawa is now considering these factors and may re-examine for the presence of aromatase in the mammalian brain.

-2- Now that Dr. Osawa has synthesized several derivitives of nor- nicotine and anabasine which are potent anti-aromatases,he is planning to do=• in vitro kinetic studies of them; to undertake in vivo assessment of their eff.ects on endocrine function and on estrogen dependent tumors in a rat model; to determine the active site of aromatase by use of tobacco aromatase inhibitors; and to determine the effects of these inhibitors during pregnancy and on the endocrine function of the offspring, as well as the effect on general growth and development. Since the effect of aromatase on male behavioural development occurs shortly after birth, he plans to inject male animals IP within 2-3 days of birth with the inhibitors. He also intends to determine if there is an effect on the female. Other questions under consideration are: Can such inhibitors of aromatase be synthesized in vivo from nicotine, or must they be obtained from smoke. Will they pass the placental barrier and will they appear in the milk. Further, do they bind to nicotine or estrogen receptors, or.is there only an action on the enzyme. Many of Dr. Osawa's recent publications have not indicated support by CTR. He explains that they were all undertaken before he started to receive support from the*Council. Two recent publications indicate our support:"Aromatase isozyme in goldfish brain and stereochemistry of the aromatization," Kitawaki, et al. (In press, Steroids) and "Immunoaffinity purification of aromatase cytochrome P-450 from human placental microsomes. Metabolic switching from aromatization to 10 and 20-monohydroxylation, and recognition of aromatase isozymes," Osawa, et al., Steroids 50:11-28, 1987. (They are a year late in publishing the journal). Comment: Dr. Osawa continues to make progress in the understanding of the chemistry of the antiaromatase in tobacco leaves or smoke. He has now arrived at the point where he is undertaking a number of in vivo studies which should extend our knowledge of how they may alter endocrine function or influence the development of estrogen- dependent cancers, as well as effects on the developing fetus. This program appear to continue to be relevent to the interests of CTR DHF