Phenyl vinyl ether의 돌연변이 활성화

Abstract

[한글]

다양한 종류의 화학 발암물질에 존재하는 ethylenic group은 생체내에서 epoxidation되어 발암성을 나타낸다. 이러한 화합물로는 aflatoxin B^^1 , vinyl chloride, vinyl carbamate, styrene 등이 있으며, 다양한 다환성 방향족 탄수화물도 여기에 속한다. 그 중 4-nitrophenyl vinyl ether(NPVE)는 cytochrome P-450의 효소 활성을 위한 기질로 사용되며 NPVE의 epoxide 대사산물인 2'-(4-nitrophenoxy)oxirane(NPO)은 물에서 불안정하여 쉽게 glycolaldehyde와 phenol로 분해된다. NPVE의 돌연변이성은 N-hydroxy NPVE에 의해 생길 가능성도 있고 2'-(4-nitrofhenoxy)oxirane에 의해 생길 가능성도 있다. 그러므로 본 연구에서는 nitro기가 없는 phenyl vinyl ether(PVE)를 합성하고, 최종 발암물질로 예상되는 phenoxyoxirane(PO)을 합성하여 이 물질의 생물화학적 성질과 돌연변이성 및 발암성을 알아보고, PVE가 microsomal oxidation 과정을 거쳐 PO로 되어 최종 친전자성 발암물질로 작용하는지를 실험하여 다음과 같은 결과를 얻었다.

PVE에 강력한 산화제인 m-chloroperoxybenzoic acid(M-CPBA) 또는 dimethyldioxirane(DMDO)을 처리하여 PO를 합성하였을 때 재래식 방법인 m-CPBA보다 DMDO를 처리하였을 때 생성률이 높았다. 순수하게 만들어진 PO의 화학적 반감기는 2.7분이었으며, 154mM chloride ion이 존재하는 경우 1.9분으로 감소되었다. 또한 PO는 물속에서 glycolaldehyde와 phenol로 분해되었으며, chloride ion 존재하에서는 이 두 화합물 이외에도 chloroacetaldehyde와 미지의 화합물이 더 검출되었다. PVE는 microsomal oxidation 과정을 거쳐 최종산물로 glycolaldehyde가 생성되었고, cytochrome P-450 억제물질인 diethyldithiocarbamate 존재하에서는 glycolaldehyde 생성이 억제되었으며, PO는 물속에서 자연 가수분해되어 최종산물로 glycolaldehyde가 생성되었다. PVE는 microsomal activation system 존재하에 약한 돌연변이성을 나타냈으나 PO는 대사 활성 과정 없이도 S.typhimurium TA98과 TA1535에 모두 강한 돌연변이성을 나타냈다. PO는 DNA와 반응하여 주 부가물로서 7-(2'-oxoethyl)guanine과 부 부가물로서 N**2 , 3-ethenoguanine을 생성하였다. 또한 PVE는 자성 ICR계 mice 피부 발암실험에서 약한 발암성을 나타냈으나 PO는 강한 발암성을 나타냈다.

이상의 결과로 보아 발암 전구물질(procarcinogen)인 PVE는 cytochrome P-450이나, 이효소를 함유한 microsome에 의해 활성화되어 최종 발암 물질인 PO로 되고, 이 물질은 glycolaldehyde나 phenol로 불활성화 되기도 하나 일부는 DNA와 반응해서 7-(2'-oxoethyl)guanine이나 N**2 , 3-ethenoguanine과 같은 DNA 부가물을 만들며, DNA수정후 복제 과정중에 유전자에 돌연변이를 유발하여 발암성을 나타내는 것으로 사료된다.

[영문]

It is well-known that ethylenic groups in a wide variety of carcinogenic compounds are the targets of epoxidation. The typical compounds containing these groups are included in aflatoxin B^^1 , various polycyclic aromatic hydrocarbons, styrene, vinyl chloride, vinyl carbamate and 4-nitrophenyl vinyl ether(NPVE). Most of NPVE epoxide, 2'-(4-nitrophenoxy)oxirane(NPO), is unstable in the water and rapidly hydrolized into glycolaldehyde and 4-nitrophenol. However, minor

proportions of the epoxides leads to mutagenic activity by formation of DNA adducts; in bacteria, the nitro residue is activated by the nitro reductase to form N-hydroxy nitro phenyl vinyl ether, whereas the epoxide residue is formed by cytochrome P-450 oxidation system in mammal. It was postulated that metabolic intermediates of NPVE, NPO or N-hydroxy nitro phenyl vinyl ether have a mutagenic activity. Therefore, two compounds of NPVE derivatives, phenyl vinyl ether(PVE) and phenoxyoxirane(PO) were synthesized in order to clarify whether the nitro group of NPVE was related to mutagenicity. PVE and PO were synthesized to test the possibilities that the mutagenic activity is related to cellular transformation. Biological properties of these two compounds were characterized and their mutagenicity and carcinogenicity were tested. The results of this study are summarized as follows.

The yield of PO production from PVE using dimethyldioxirane or

m-chloroperoxybenzoic acid was 98% and 80%, half lives of PO in the absence or presence of 154 mM chloride ion at 37。C were 2.7 min and 1.9 min, respectively. PO was rapidly degraded into phenol and glycolaldehydein the water spontaneously. However, it was more rapidly degraded in the presence of chloride ion and additional chloroacetaldehyde and some unknown products were formed. Glycolaldehydeyde also was formed as an end product of the microsomal oxidation of PVE, and diethyldithiocarbamate, a cytochrome P-450 inhibitor, inhibited the formation of glycolaldehyde. PO was strong mutagenic in S. typhimurium TA98 and TA1535, while its precursor PVE was weak mutagenic in the presence of P-450 microsomal activation system. DNA adduct formation of PO, 7-(2'-oxoethyl)guanine or N**2,3-ethenoguanine was dose dependent in vitro. PO was strongly carcinogenic molecule in the female ICR mouse skin while its precursor PVE was weak carcinogenic.

From these results, it is suggested that PVE is metabolized into PO by microsomal oxidation and the PO reacted with DNA to form DNA adducts, which leads to mutagenic and carcinogenic activities by the introduction of DNA mutations in the genes

responsible for cellular transformation in the course of repairing DNA adducts by the host cellular process.