Probing Bioactive Mechanisms - American Chemical Society

Chapter 22

Structurally Specific Interaction of Halogenated Dioxin and Biphenyl Derivatives with Iodothyronine-5'-deiodinase in Rat Liver U. Rickenbacher , S. Jordan, and J . D. McKinney 1

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Downloaded by UNIV OF AUCKLAND on April 2, 2018 | https://pubs.acs.org Publication Date: November 14, 1989 | doi: 10.1021/bk-1989-0413.ch022

Laboratory of Molecular Biophysics, National Institute of Environmental Health Sciences, P.O. Box 12233, Research Triangle Park, NC 27709 In in vitro studies, soluble, polar derivatives of polychlorinated biphenyls (PCB) and dibenzo-p-dioxins were shown to inhibit outer (phenolic) ring deiodination of 3,3',5'-triiodothyronine (rT3) used as substrate for iodothyronine type I deiodinase activity in microsomal fractions of rat liver. Potent inhibition depended on the presence of lateral chlorination (3,5 in biphenyl or 2,3 in dibenzo-p-dioxin derivatives). The most potent PCB ligand exhibited a half-maximal inhibitory concentration similar to the Km (29 nM) of rT3. The results are in general agreement with our previous results with human thyroxine binding prealbumin and rat liver nuclear extracts that also show high affinity specific binding of these and related compounds to thyroxine specific binding sites. The functional structural characteristics of these polar PCB and dioxin derivatives involved in binding are in general similar to those found in toxic structures (underivatized) of this type. These relatively metabolically resistant deiodination inhibitor analogs may be useful as selective inhibitors facilitating the further study of biochemical and functional characteristics of protein interactions in thyroid hormone metabolism as well as the study of the possible importance of thyroid hormone antagonism in dioxin and related compound toxicity. The t o x i c p o t e n c y o f h a l o g e n a t e d a r o m a t i c h y d r o c a r b o n s i s dependent on t h e number and p o s i t i o n s o f h a l o g e n atoms i n t h e i r m o l e c u l a r structure. Q u a l i t a t i v e s t r u c t u r e requirements f o r h i g h t o x i c i t y i n c l u d e p l a n a r i t y ( o r c o p l a n a r i t y ) o f s t r u c t u r e i n a shape a p p r o x i m a t i n g a r e c t a n g l e and a s u f f i c i e n t degree o f h a l o g e n a t i o n Current address: Sandoz Limited 881, C H 4002, Basel, Switzerland Address correspondence to this author.

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This chapter not subject to U.S. copyright Published 1989 American Chemical Society

Magee et al.; Probing Bioactive Mechanisms ACS Symposium Series; American Chemical Society: Washington, DC, 1989.

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a p p r o x i m a t i n g a r e c t a n g l e and a s u f f i c i e n t degree o f h a l o g e n a t i o n concentrated about l a t e r a l p o s i t i o n s ( 3 , 3 , 4 , 4 ' , 5 , 5 * - p o s i t i o n s on the b i p h e n y l n u c l e u s and 2,3,7,8-positions on t h e d i o x i n n u c l e u s ) o f the molecule (I). Treatment o f animals w i t h 2 , 3 , 7 , 8 - t e t r a c h l o r o d i b e n z o - p - d i o x i n (TCDD), t h e p r o t o t y p e f o r c e r t a i n t o x i c halogenated aromatic hydrocarbons, produces divergent e f f e c t s on c i r c u l a t i n g l e v e l s o f L - t h y r o x i n e ( L - T , ) and L - 3 , 5 , 3 ' - t r i i o d o t h y r o n i n e (L-Tg) concentrations (2-5). These r e s u l t s a l o n g w i t h t h e l a c k o f i n f o r m a t i o n on t h y r o i d hormone modulated r e s p o n s e s i n t h e s e a n i m a l s have caused u n c e r t a i n t y r e g a r d i n g t h e i r f u n c t i o n a l and b i o c h e m i c a l t h y r o i d s t a t u s ( 4 ) . B e c a u s e o f t h e s t r u c t u r a l s i m i l a r i t i e s ( 6 ) b e t w e e n TCDD a n d T a n d t h e f a c t t h a t t h y r o i d h o r m o n e s c a n m o d u l a t e TCDD t o x i c i t y ( 7 - 8 ) , we f o c u s e d o u r a t t e n t i o n o n t h y r o x i n e b i n d i n g p r o t e i n s a s p o s s i b l e s i t e s f o r c r i t i c a l b i o c h e m i c a l i n t e r a c t i o n s t h a t may m e d i a t e t h e t o x i c r e s p o n s e s o f t h e s e compounds. Initial studies ( 9 - 1 0 ) i n v o l v e d human t h y r o x i n e b i n d i n g p r e a l b u m i n ( t r a n s t h y r e t i n o r TBPA) ( 9 - 1 0 ) s i n c e i t i s a major t r a n s p o r t p r o t e i n f o r T^ i n b l o o d a n d may be o f u s e a s a m o d e l f o r s t u d y i n g t h e i n t e r a c t i o n o f t h y r o i d hormones w i t h c e r t a i n n u c l e a r t h y r o x i n e r e c e p t o r s (1_0. C o m p e t i t i v e b i n d i n g s t u d i e s u s i n g TBPA a n d t h e t h y r o x i n e n u c l e a r r e c e p t o r e x t r a c t e d f r o m r a t l i v e r (12) a n d p o l a r ( s o l u b l e ) d e r i v a t i v e s o f d i o x i n and r e l a t e d p o l y c h l o r i n a t e d b i p h e n y l ( P C B ) compounds have shown t h a t s i m p l e h a l o g e n a t e d h y d r o c a r b o n s w h i c h c o n t a i n o n l y one aromatic ring or l i n e a r structures with multiple rings bind with higher a f f i n i t y than the normal angular d i p h e n y l e t h e r bridged system c h a r a c t e r i s t i c o f t h y r o i d hormone a n a l o g s . Furthermore, l a t e r a l c h l o r i n a t i o n w a s common t o a l l c o m p o u n d s w h i c h s h o w e d h i g h b i n d i n g activity. I n a d d i t i o n , t h e n u c l e a r r e c e p t o r showed a r e m a r k a b l y enhanced a f f i n i t y f o r l a t e r a l l y s u b s t i t u t e d compounds t h a t were a l s o p l a n a r and h i g h l y p o l a r i z a b l e . Because these s t r u c t u r a l properties are a l s o c h a r a c t e r i s t i c o f h i g h l y t o x i c s t r u c t u r e s o f t h i s type, i t was s u g g e s t e d ( 6 ) t h a t t o x i c i t y may be i n p a r t t h e e x p r e s s i o n o f p o t e n t and p e r s i s t e n t t h y r o i d hormone a g o n i s t a c t i v i t y i n i t i a t e d a t the l e v e l o f the nuclear receptor. In contrast, c e r t a i n nonreceptor t h y r o i d h o r m o n e b i n d i n g p r o t e i n i n t e r a c t i o n s may be a n t a g o n i s t i c i n nature.


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P r e v i o u s w o r k e r s (13) h a v e s h o w n s t r o n g c o r r e l a t i o n s b e t w e e n t h e b i n d i n g r e q u i r e m e n t s f o r TBPA a n d t h e i n h i b i t o r y r e q u i r e m e n t s f o r rat l i v e r iodothyronine deiodinase a c t i v i t y . The i o d o t h y r o n i n e raonodeiodinases (ITHD) a r e t h e k e y enzymes o f e x t r a t h y r o i d a l t h y r o x i n e m e t a b o l i s m (14-15). Inactivation of thyroxine's t h y r o m i m e t i c p o t e n c y o c c u r s by t y r o s y l 5 - d e i o d i n a t i o n , p r o d u c i n g reverse-T (rTg). r T ( 3 , 3 5 ' - t r i i o d o t h y r o n i n e ) i s t h e most p o t e n t o f t h e p h y s i o l o g i c a l l y o c c u r r i n g i n h i b i t o r s o f ITHD a n d i s t h o u g h t t o be o f r e g u l a t o r y i m p o r t a n c e i n m o d u l a t i n g d e i o d i n a t i v e iodothyronine metabolism (14-15). Previous studies suggest the e x i s t e n c e o f a t l e a s t t h r e e ITHD i s o z y m e s ( T y p e s I - I I I ) w i t h d i f f e r e n t organ d i s t r i b u t i o n and f u n c t i o n i n T m e t a b o l i s m ( 1 4 - 1 5 ) . The b e s t c h a r a c t e r i z e d i s o z y m e , t y p e I ITHD, p r e f e r s r T ( K 5 0 - 1 0 0 nM) o v e r T (K - 2 pM) a s a s u b s t r a t e f o r d e i o d i n a f S o n , r e q u i r e s reduced d i t h T o f i ^ a s c o s u b s t r a t e s , and r e a c t s i n a p i n g - p o n g mechanism. T y p e I ITHD i s p r i m a r i l y a s s o c i a t e d w i t h t h e m i c r o s o m a l 3

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f r a c t i o n o f l i v e r , k i d n e y , and t h e e u t h y r o i d b r a i n and i s r e s p o n s i b l e f o r t h e p r o d u c t i o n o f a b o u t 70% o f t h e c i r c u l a t i n g T i n the e u t h y r o i d r a t (14-15). Thus, the p o t e n t i a l i n h i b i t o r y e f f e c t s o f t o x i c h a l o g e n a t e d a r o m a t i c h y d r o c a r b o n s o n t h e t y p e I ITHD isozyme were o f i n t e r e s t s i n c e such e f f e c t s c o u l d modulate t h y r o i d hormone m e t a b o l i s m and b i o c h e m i c a l t h y r o i d s t a t u s . Previous studies have shown t h a t s u c h d i v e r s e c l a s s e s o f compounds a s i o d i n a t e d contrast media, i n d i c a t o r dyes, anticoagulants, f l a v o n o i d s , plant e x t r a c t s , c a t e c h o l a m i n e ( a n t ) a g o n i s t s , and t h i o u r e a d e r i v a t i v e s a c t a s p o t e n t i n h i b i t o r s o f t y p e I ITHD ( 1 3 - 1 5 ) . I n t h i s w o r k , we investigated the i n v i t r o deiodinase i n h i b i t o r y a c t i v i t y of a s e l e c t e d number o f h y d r o x y l a t e d PCBs a n d s o l u b l e s t r u c t u r a l p r o b e s f o r d i o x i n s t h a t had p r e v i o u s l y been shown t o have d i s t i n c t b i n d i n g a f f i n i t i e s t o TBPA ( 9 - 1 0 ) . Experimental

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Compounds. r - T f r e e a c i d was p u r c h a s e d from Sigma C h e m i c a l C o . ( S t . L o u i s , MO). H y d r o x y l a t e d P C B s w e r e o b t a i n e d f r o m U l t r a S c i e n t i f i c (Hope, R I ) w i t h t h e e x c e p t i o n o f TCDB-2 ( s e e T a b l e 1 f o r d e f i n i t i o n o f a b b r e v i a t i o n s u s e d ) w h i c h was s y n t h e s i z e d p r e v i o u s l y i n o u r l a b o r a t o r y by a d a p t a t i o n o f methods i n t h e PCB s y n t h e t i c l i t e r a t u r e ( L 6 ) . T r i C D D A a n d DDA w e r e s y n t h e s i z e d i n o u r l a b o r a t o r y as d e s c r i b e d e a r l i e r ( 9 ) . [ I ] r-T ( s p e c i f i c a c t i v i t y 1000-1400 y C i / j i g ) w a s p u r c h a s e d f r o m New E n g l a n d N u c l e a r C o r p . (Boston, MA). If necessary [ I ] r - T ^ was p u r i f i e d b e f o r e u s e by p a p e r electrophoresis (17). 3

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Animals. M a l e S p r a g u e - D a w l e y r a t s ( C h a r l e s R i v e r , MO) w e i g h i n g 150 t o 300g were u s e d . T h e y w e r e f e d N I H 31 d i e t a n d h o u s e d i n r o o m s k e p t a t 21 ± i C a n d 50% r e l a t i v e h u m i d i t y , w i t h a 1 2 - h r l i g h t / d a r k c y c l e ( l i g h t e d f r o m 0700 t o 1900 h r ) . The i n v i t r o enzyme i n h i b i t i o n s t u d i e s were performed u s i n g combined l i v e r m i c r o s o m a l p r e p a r a t i o n s from 6 r a t s as f e d and housed i n t h i s manner. M i c r o s o m a l p r e p a r a t i o n s were o b t a i n e d i n t h e f o l l o w i n g way. L i v e r w a s r e m o v e d , q u i c k l y c h i l l e d i n c o l d b u f f e r ( 2 0 mM t r i s ( h y d r o x y m e t h y l ) a m i n o m e t h a n e ( T r i s ) , 0 . 3 2 M s u c r o s e pH 7 . 0 ) and b l o t t e d on f i l t e r paper. A p p r o x i m a t e l y 3 g were minced and homogenized i n 6 volumes o f b u f f e r w i t h a motor d r i v e n t e f l o n pestle. T h e h o m o g e n a t e w a s c e n t r i f u g e d f o r 15 m i n a t 1 8 , 0 0 0 x g . The p e l l e t was r e s u s p e n d e d i n 7 m l b u f f e r a n d c e n t r i f u g e d a s a b o v e . The c o m b i n e d s u p e r n a t a n t was c e n t r i f u g e d f o r 1 h a t 1 0 0 , 0 0 0 x g . The m i c r o s o m a l p e l l e t was m i x e d w i t h 0 . 2 M p o t a s s i u m p h o s p h a t e b u f f e r ( p H 7 . 0 ) c o n t a i n i n g 1 mM e t h y l e n e d i a m i n e t e t r a a c e t a t e (EDTA) and was homogenized i n 8 m l o f t h i s b u f f e r . An a l i q u o t f o r the p r o t e i n d e t e r m i n a t i o n w a s s a v e d a n d t h e r e m a i n d e r made 1 mM i n d i t h i o t h r e i t o l (DTT) and f r o z e n i n d r y i c e - a c e t o n e . The samples w e r e s t o r e d a t - 7 0 ° C f o r n o l o n g e r t h a n 45 d a y s . The p r o t e i n c o n c e n t r a t i o n o f t h e D T T - f r e e m i c r o s o m a l p r e p a r a t i o n was d e t e r m i n e d (18) u s i n g b o v i n e serum a l b u m i n as a s t a n d a r d . #

rT3 5 ' - d e i o d i n a s e a s s a y . m i c r o s o m e s was d e t e r m i n e d

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[ I ] - r T ^ ( w i t h m o d i f i c a t i o n s ) based on t h e p r e v i o u s method ( 1 9 ) . In t h i s assay rT^ i s used as s u b s t r a t e i n s t e a d o f T^ because T^ a l l o w s a l t e r n a t e ( i n n e r r i n g ) d e i o d i n a t i o n c o m p l i c a t i n g t h e s p e c i f i c measurement o f o u t e r r i n g d e i o d i n a t i o n . Direct in vitro i n h i b i t i o n o f t h e 5 ' - d e i o d i n a s e by h y d r o x y l a t e d PCBs was a n a l y z e d as follows: I n a t o t a l v o l u m e o f 200 u l t h e r e a c t i o n m i x t u r e c o n t a i n e d 0 . 2 M p o t a s s i u m p h o s p h a t e , 1 mM D T T , 1 mM E D T A , 10 u g m i c r o s o m a l p r o t e i n a n d 1 nM [ I ] r T ( p H 7 . 0 ) . The s p e c i f i c a c t i v i t y was a p p r o x i m a t e l y 5 2 0 , 0 0 0 ^ c p m / p m o | r T - . The PCBs were added i n 5 u l m e t h a n o l y i e l d i n g 10~~ t o 1 0 " M i n t h e r e a c t i o n m i x t u r e . Reaction was s t a r t e d b y t h e a d d i t i o n o f t h e m i c r o s o m e s . I n c u b a t i o n was c a r r i e d o u t a t 3 7 C f o r 5 m i n . The r e a c t i o n was t e r m i n a t e d b y t h e a d d i t i o n o f 3 0 0 u l o f i c e - c o l d s o l u t i o n o f 10 uM L - T a n d 5 0 uM 6 - p r o p y l - 2 - t h i o u r a c i l (PTU). L i b e r a t e d I~ w a s q u a n t i f i e d b y s e p a r a t i n g I~ f r o m r T o n d i s p o s a b l e S e p h a d e x G - 2 5 m i n i c o l u m n s a n d subsequent counting of [ I ] i n a P a c k a r d A u t o Gamma c o u n t e r (60% e f f i c i e n c y ) as d e s c r i b e d e a r l i e r (TO). B r i e f l y , a f t e r the r e a c t i o n was s t o p p e d a 400 u l a l i q u o t was a d d e d t o t h e c o l u m n a n d e l u t e d w i t h 1.3 m l 0 . 2 M p o t a s s i u m p h o s p h a t e b u f f e r ( p H 7 . 0 ) . T h i s f i r s t fraction contained [ I ] r T ^ e v e n t u a l l y bound t o p r o t e i n s ( t y p i c a l l y < 2% o f t o t a l s u b s t r a t e c o n c e n t r a t i o n ) . The f i r s t f r a c t i o n was s e p a r a t e d f r o m t h e s e c o n d f r a c t i o n (2 m l ) c o n t a i n i n g [ I ]. Free r T ^ b i n d s t i g h t l y t o t h e g e l and does n o t e l u t e i n the volume used. To c o r r e c t f o r n o n e n z y m a t i c d e i o d i n a t i o n and contamination with [ 1 ~ ] , c o n t r o l i n c u b a t i o n s were conducted w i t h o u t m i c r o s o m e s o r w i t h b o i l e d m i c r o s o m e s y i e l d i n g t h e same r e s u l t (1 t o 21 o f t o t a l c o u n t s ) . The m o l a r amount o f i " l i b e r a t e d was c a l c u l a t e d by r e f e r r i n g t o h a l f o f t h e s p e c i f i c a c t i v i t y f o r r T g a s s u m i n g a n e q u a l c h a n c e f o r t h e 3 o r 5 p o s i t i o n s t o be o c c u p i e d by [ I ] (19). Because o f s o l u b i l i t y problems the a c t i v i t y o f T r i C D D A a n d DDA w a s m e a s u r e d i n a m o d i f i e d p r o c e d u r e : T o t a l v o l u m e was 500 u l ( o p p o s e d t o 200 u l ) i m i d a z o l e b u f f e r ( p H 7 . 0 ) w i t h 5 u g ( o p p o s e d t o 10 u g ) m i c r o s o m a l p r o t e i n , 0 . 6 nM [ I]rT ( 1 nM t o t a l r T ) a n d 5 mM D D T . T h e e n z y m e r e a c t i o n w a s much s l o w e r u n d e r t h e s e c o n d i t i o n s a n d l i n e a r u p t o 60 m i n . A n i n c u b a t i o n t i m e o f 45 m i n w a s c h o s e n . A t t h i s t i m e p o i n t a g a i n about 201 o f t h e s u b s t r a t e was c o n v e r t e d a t t h e h i g h e s t r a t e . Reaction was t e r m i n a t e d by q u i c k l y c o o l i n g t h e s a m p l e s o n i c e a n d a d d i n g them d i r e c t l y onto the columns. The t i m e f r o m t h e t e r m i n a t i o n o f t h e enzyme r e a c t i o n t o t h e c o m p l e t e e l u t i o n o f t h e s a m p l e s t o o k l e s s than 2 min f o r each sample. 1 2 ,

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The L i n e w e a v e r - B u r k ( 2 0 ) p l o t f o r t h e i n h i b i t o r i n t e r a c t i o n s was o b t a i n e d i n t h e f o l l o w i n g w a y . T h e r e a c t i o n m i x t u r e c o n t a i n e d 10 u g m i c r o s o m a l p r o t e i n , 0 . 0 5 nM - 0 . 6 nM [ I]-rT [total rT c o n c e n t r a t i o n w a s 5 t o 6 0 nM i n 2 0 0 u l o f 0 . 2 M p o t a s s i u m p h o s p h a t e b u f f e r ( 1 mM D T T , 1 mM E D T A , p H 7 . 0 ) ] a l o n e o r i n t h e p r e s e n c e o f 1.5 uM T C H B - 1 , 0 . 1 5 uM T C D B - 2 , 6 . 2 5 uM T r i C D D A o r 0 . 1 1 uM T C D B - 1 . H y d r o x y l a t e d PCBs and r T were added i n 5 u l m e t h a n o l and t h e d i o x i n d e r i v a t i v e s i n 5 u l d i r a e t h y l s u l f o x i d e (DMSO). There was no d i f f e r e n c e i n t h e o f r T ( 2 9 nM) b e t w e e n t h e m e t h a n o l a n d t h e DMSO c o n t a i n i n g r e a c t i o n . I n c u b a t i o n t i m e was 5 m i n a t 3 7 * C R e s u l t s a r e t h e mean o f d u p l i c a t e d e t e r m i n a t i o n s a n d r e p r e s e n t a t i v e o f two o r more i n d e p e n d e n t e x p e r i m e n t s . Linear regression analysis gave c o r r e l a t i o n c o e f f i c i e n t s > 0 . 9 f o r a l l the l i n e s . 1 2 ,

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PROBING BIOAC1TVE MECHANISMS

Results Type I d e i o d i n a s e a c t i v i t y i s p r i m a r i l y a s s o c i a t e d w i t h the microsomal f r a c t i o n of l i v e r tissue (14-15). The d e i o d i n a s e a c t i v i t y was a s s a y e d u s i n g c o n d i t i o n s a d a p t e d f r o m t h e literature ( 1 9 ) b u t u s i n g r e l a t i v e l y more d i l u t e c o n d i t i o n s t o e n h a n c e t h e b i n d i n g a c t i v i t y and f a c i l i t a t e s t u d y o f t h e more l i p o p h i l i c c h l o r i n a t e d aromatic hydrocarbon d e r i v a t i v e s . The enzyme r e a c t i o n was l i n e a r w i t h i n c u b a t i o n t i m e and p r o t e i n c o n c e n t r a t i o n f o r a l l c o n d i t i o n s d e s c r i b e d i n the E x p e r i m e n t a l S e c t i o n . In a d d i t i o n , the s u b s t r a t e was n e v e r d e p l e t e d b y m o r e t h a n a b o u t 20%. For example, i n the assay used f o r the h y d r o x y l a t e d PCBs, 10.5 fmols of r T was c o n s u m e d i n 1 rain ( 5 . 3 % o f t o t a l s u b s t r a t e ) , 2 8 . 2 f m o l s a t 3 m i n ( 1 4 . 1 % ) , and 4 7 . 6 f m o l s a t 6 m i n ( 2 3 . 8 % ) . T h e same was t r u e o f t h e p r o c e d u r e used f o r the d i o x i n d e r i v a t i v e s ( d a t a n o t shown), and u n d e r t h e s e c o n d i t i o n s t h e r e a c t i o n was m u c h s l o w e r ( l i n e a r u p t o 60 min)(Figure 1).


3

C o m p e t i t i v e i n h i b i t i o n s t u d i e s were done u s i n g s e l e c t e d p o l a r h y d r o x y l a t e d PCB a n d a d i p a m i d e d i o x i n d e r i v a t i v e s a v a i l a b l e f r o m p r e v i o u s w o r k w i t h TBPA ( 9 - 1 0 ) . F i g u r e s 2 and 3 show t h e e f f e c t o f i n c r e a s i n g c o n c e n t r a t i o n s o f v a r i o u s h y d r o x y l a t e d PCBs on t h e c o n v e r s i o n of rT^ i n t o 3 , 3 - d i i o d o t h y r o n i n e ( T ^ ) . From these p l o t s , i n h i b i t o r y concentrations y i e l d i n g half-maximal i n h i b i t i o n (IC 's) a r e e s t i m a t e d (1_3). A c o m p a r i s o n o f t h e s e v a l u e s i s s h o w n i n T a b l e 1. L a t e r a l c h l o r i n e s u b s t i t u t i o n ( 3 , 5 o r m e t a ) was common t o t h e t w o m o s t a c t i v e P C B c o m p o u n d s (TCDB 1 a n d 2 ) . The i n a c t i v i t y o f T C H B - 2 a n d DDA w h i c h l a c k l a t e r a l s u b s t i t u e n t s ( 3 , 5 o n b i p h e n y l o r 2,3 on d i o x i n n u c l e u s ) f u r t h e r s u p p o r t s the i m p o r t a n c e o f l a t e r a l substitution for significant binding a c t i v i t y . The r e s u l t w i t h DDA a l s o argues a g a i n s t the p o s s i b i l i t y t h a t the adipamide group i s a dominant f a c t o r i n b i n d i n g . The r e l a t i v e l y l e s s p o l a r a n d m o r e l i p o p h i l i c compounds, TCHB-1 and T r i C D D A showed c o n s i d e r a b l y l o w e r inhibitory activity. However, the s i m i l a r i n h i b i t o r y potency of t h e s e t w o c o m p o u n d s w o u l d s u g g e s t t h a t a l a t e r a l h y d r o x y l g r o u p may n o t be a c r i t i c a l f a c t o r i n b i n d i n g . 1

Table

1

IC^- values o f c h l o r i n a t e d b i p h e n y l and d i o x i n i n the 5 ' - d e i o d i n a t i o n of r T . Inhibitor

Abbrev.

3,3',5,5'-tetrachlorodihydroxybiphenyl 2,3,5,6-tetrachlorodihydroxybiphenyl 3 , 7 , 8 - t r i c h l o r o d i b e n z o d i o x i n adipamide 3,5,4'-trichloro-4-hydroxybiphenyl 2,4,6-trichloro-4'-hydroxybiphenyl d i b e n z o d i o x i n adipamide

TCDB-1 TCDB-2 TriCDDA TCHB-I TCHB-2 DDA

derivatives

IC

5 Q

0.07 0.32 4.40 7.20 N A

b NA

^ H a l f - m a x i m a l c o n c e n t r a t i o n s e s t i m a t e d from F i g s . 2 and 3. No a p p r e c i a b l e a c t i v i t y was o b s e r v e d a t t h e c o n c e n t r a t i o n o f inhibitor tested.


Probing Bioactive Mechanisms - American Chemical Society

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