Europaisches Patentamt
`European Patent Office
`Office europeen des brevets
`
`© Publication number:
`
`0 1 8 1
`A 2
`
`5 9 2
`
`©
`
`EUROPEAN PATENT APPLICATION
`
`© Application number: 85114042.6
`© Date of filing: 05.11.85
`
`© Int. CI.4: C 07 C 1 2 9 / 1 2
`A 61 K 3 1 / 1 6
`
`Go) Priority: 13.11.84 JP 237480/84
`
`@ Date of publication of application:
`21.05.86 Bulletin 86/21
`© Designated Contracting States:
`AT BE CH DE FR GB IT LI LU NL SE
`© Applicant: MICROBIAL CHEMISTRY RESEARCH
`FOUNDATION
`14-23, Kamiosaki 3 Chome Shinagawa-ku
`Tokyo 141 (JP)
`© Inventor: Umezawa, Hamao, Prof.
`23 Toyotama-kha 4-chome
`Nerima-ku Tokyo(JP)
`© Inventor: Takeuchi, Tomio
`1-11, Higashigotanda 5-chome
`Shinagawa-ku Tokyo(JP)
`
`© Inventor: Ishizuka, Masaaki
`17, Denenchofu-honcho 3 chome
`Ohta-kuTokyo(JP)
`© Inventor: Abe, Fuminori
`29-15, Shimo 3 -chome
`Kita-ku Tokyo! JP)
`@ Inventor: Fujii.Akio
`8-13, Of una 4-chome
`Kamakura-cfty Kanagawa Prefecture! JP)
`© Inventor: Nakamura, Teruya
`831, Nomura-cho
`Kusatsu-city Shiga Prefecture! JP)
`© Representative: Becker, Heinrich Karl Engelbert,
`Dr. et at,
`HOECHST AKTIENGESELLSCHAFT Central Patent
`Department P.O. Box 80 03 20
`D-6230 Frankfurt am Main 80(DE)
`
`•
`
`(I)
`
`@ Use of spergualin derivatives for the preparation of medicaments having immunosuppressive activity.
`© The present invention relates to the use of compounds
` T h e present invention relates to the use of compounds
`of the formula
`of the formula
`NH
`II
`NHj-C-NH-R,-R2-CONH-R3-CONH-(CH2)4-NH-(CH2)3-NH2
`wherein R, is -(CH3)4-,-(CH2)6-, -^^-CH2' "^^"(CH2)2~
`'• R2 is HCH2)2- or -CH=CH-; and R3 is -CH-,
`OH
`
`or
`
`-CHg-^C^-
`-CH„
`
`-CH-, -CH2- or -CH-
`I
`I
`CH2OH
`OCH3
`or pharmacologically acceptable salts thereof, for the prepa-
`or pharmacologically acceptable salts thereof, for the prepa-
`ration of medicaments having immunosuppressive activity.
`ration of medicaments having immunosuppressive activity.
`
`Croydon Printing Company Ltd
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 1 of 25
`
`

`
`r e l a t e s
`to
`i n v e n t i o n
`The p r e s e n t
`i m m u n o s u p p r e s s a n t s
`c o n t a i n i n g S p e r g u a l i n compounds as an a c t i v e
`i n g r e d i e n t .
`i m m u n o s u p p r e s s i v e a c t i o n s a r e
`A number of d r u g s w i t h
`i n c l u d e a l k y l a t i n g a g e n t s , a n t i m e t a b o -
`a l r e a d y known. They
`f o l i c a c i d a n t a g o n i s t s a n d
`s t e r o i d s ,
`l i t e s , a n t i b i o t i c s ,
`p l a n t a l k a l o i d s .
`i s o l a t e d by Umezawa
`t h a t was
`is a compound
`S p e r g u a l i n
`t h e f i l t r a t e of a c u l t u r e b r o t h of S p e r g u -
`and o t h e r s from
`a l i n - p r o d u c i n g m i c r o o r g a n i s m of
`the g e n u s B a c i l l u s . T h e
`s t r u c t u r a l f o r m u l a of S p e r g u a l i n
`is shown b e l o w :
`
`is not o n l y e f f e c t i v e a g a i n s t mouse
`l e u k e m i a
`S p e r g u a l i n
`l e u k e m i a E L - 4 , E h r l i c h c a r c i n o m a and s a r c o m a
`L - 1 2 1 0 , mouse
`180 bus a l s o e x h i b i t s p r o m i s i n g a c t i v i t y
`in c o n t r o l l i n g m a -
`( s e e J a p a n e s e U n e x a m i n e d P u b l i s h e d P a t e n t
`t u m o r s
`l i g n a n t
`A p p l i c a t i o n No. 4 8 9 5 7 / 1 9 8 2 ) .
`Umezawa et a l . c o n t i n u e d
`t h e i r s t u d i e s on S p e r g u a l i n
`c o m p o u n d s and h a v e s y n t h e s i z e d n u m e r o u s S p e r g u a l i n c o m -
`p o u n d s which a r e more s t a b l e and have a s t r o n g e r a n t i -
`( s e e J a p a n e s e U n e x a m i n e d P u b l i s h e d P a t e n t
`t u m o r a c t i v i t y
`A p p l i c a t i o n Nos. 6 2 1 5 2 / 1 9 8 3 , 4 2 3 5 6 / 1 9 8 4 and 7 6 0 4 6 / 1 9 8 4 ) .
`t h o s e used
`t h e s e c o m p o u n d s are
`in
`the p r e s e n t
`i n v e n -
`Among
`t i o n .
`i m m u n o s u p p r e s s i v e e f f e c t s of s t e r o i d s a r e c o n -
`The
`to be a c c o m p l i s h e d p r i n c i p a l l y
`s i d e r e d
`the a n t i -
`t h r o u g h
`i n f l a m m a t o r y a c t i o n and
`the
`l y s i s of
`l y m p h o c y t e s . H o w e v e r ,
`s t e r o i d s have d i v e r s i f i e d p h a r m a c o l o g i c a l
`is w e l l known,
`as
`e f f e c t s and c a u s e many s i d e e f f e c t s . The o t h e r
`i m m u n o -
`s u p p r e s s a n t s a r e c l a s s i f i e d as " c y t o t o x i c " s u b s t a n c e s
`is d i r e c t e d , among o t h e r
`and
`t h e i r a c t i o n
`t h e
`to
`t h i n g s ,
`p a t h w a y s of n u c l e i c a c i d s y n t h e s i s and may o f t e n c a u s e
`s e r i o u s s i d e e f f e c t s on h e m a t o p o i e t i c o r g a n s .
`I t
`is
`t h e r e -
`f o r e d e s i r e d
`to d e v e l o p
`t h a t a c t
`i m m u n o s u p p r e s s i v e d r u g s
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 2 of 25
`
`

`
`l y m p h o c y t e s and o t h e r c e l l s of
`s e l e c t i v e l y on
`i m p o r t a n c e w h i l e c a u s i n g minimum s i d e e f f e c t s .
`i n v e n t i o n r e l a t e s
`to
`The p r e s e n t
`i m m u n o s u p p r e s s a n t s
`the S p e r g u a l i n c o m p o u n d s
`c o n t a i n i n g as an a c t i v e
`i n g r e d i e n t
`of f o r m u l a
`shown b e l o w
`( s u c h compounds a r e h e r e i n a f t e r
`(I)
`r e f e r r e d
`the c o m p o u n d s of
`i n v e n -
`the p r e s e n t
`to as
`s i m p l y
`t i o n ) :
`
`i m m u n o l o g i c a l
`
`w h e r e i n R1
`
`o r
`
`; R2
`
`is - ( C H 2 ) 2 - or -CH=CH-; and R3
`
`i s
`
`or p h a r a m c o l o g i c a l l y a c c e p t a b l e s a l t s
`t h e r e o f .
`P r e f e r r e d c o m p o u n d s a r e
`t h o s e w h e r e i n R1
`is - ( C H 2 ) 2 - ;
`or - ( C H 2 ) 6 - ; R2
`( S )
`
`is - ( C H 2 ) 4 -
`
`and R3
`
`i s
`
`t h o s e w h e r e i n R1
`
`R2
`
`is - ( C H 2 ) 2 - ; and R3
`
`is -CH2- o r
`
`t h o s e w h e r e i n R1
`and
`( t r a n s ) and R3
`i s
`
`is - ( C H 2 ) 4 - or - ( C H 2 ) 6 - ; R2
`
`is -CH=CH-
`
`the c o m p o u n d s u s a b l e as
`S p e c i f i c e x a m p l e s of
`in a c c o r d a n c e w i t h
`the p r e s e n t
`s u p p r e s s i v e a g e n t s
`l i s t e d b e l o w :
`t i o n a r e
`
`i m m u n o -
`i n v e n -
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 3 of 25
`
`

`
`(3)
`
`(5)
`
`(7)
`
`(1)
`
`N - ( 4 - ( 3 - a m i n o p r o p y l ) a m i n o b u t y l ] - 2 - ( 7 - g u a n i d i n o h e p t a n e -
`a m i d o ) - 2 - h y d r o x y e t h a n e a m i d e ;
`(2) N - [ 4 - ( 3 - a m l n o p r o p y l ) a m i n o b u t y l ] - 2 - ( 7 - g u a n i d i n o h e p t a n e -
`a m i n o ) - 2 - m e t h o x y e t h a n e a m i d e ;
`N - [ 4 - ( 3 - a m l n o p r o p y l ) a m i n o b u t y l ] - 2 - ( 9 - g u a n l d i n o n o n a n e -
`a m i d o ) - 2 - h y d r o x y e t h a n e a m i d e ;
`(4) N - [ 4 - ( 3 - a m i n o p r o p y l ) a m i n o b u t y l ] - 2 - ( 7 - g u a n i d i n o h e p t a n e -
`a m i d o ) - e t h a n e a m i d e ;
`N - [ 4 - ( 3 - a m i n o p r o p y l ) a m i n o b u t y l j - 2 - ( 7 - g u a n i d i n o h e p t a n e -
`a m i d o ) - ( S ) - 2 - h y d r o x y m e t h y l e t h a n e a m i d e ;
`(6) N - [ 4 - ( 3 - a m i n o p r o p y l ) a m i n o b u t y l ] - 2 - [ 4 - ( p - g u a n i d i n o -
`p h e n y l ) b u t a n e a m i d o ] e t h a n e a m i d e ;
`N - [ 4 - ( 3 - a m i n o p r o p y l ) a m i n o b u t y l ] - 2 - [ 4 - ( p - g u a n l d i n o -
`p h e n y l ) - b u t a n e a m i d o ] - ( S ) - 2 - h y d r o x y m e t h y l e t h a n e a m i d e ;
`(8) N - [ 4 - ( 3 - a m i n o p r o p y l ) a m l n o b u t y l ] - 2 - [ 3 - ( p - g u a n l d l n o -
`m e t h y l p h e n y l ) p r o p a n e a m i d o J - ( S ) - 2 - h y d r o x y m e t h y l e t h a n e -
`a m i d e ;
`(9) N - [ 4 - ( 3 - a m i n o p r o p y l ) a m i n o b u t y l ] - 2 - [ 5 - ( p - g u a n i d i n o -
`p h e n y l ) - p e n t a n e a m i d o J - ( S ) - 2 - h y d r o x y m e t h y l e t h a n e a m i d e ;
`(10) N - [ 4 - ( 3 - a m l n o p r o p y l ) a m i n o b u t y l ] - 2 - [ 7 - g u a n l d l n o h e p t a -
`2 - e n a m i d o ] - 2 - m e t h o x y e t h a n e a m i d e ; a n d
`(11) N - [ 4 - ( 3 - a m i n o p r o p y l ) a m i n o b u t y l ] - 2 - [ 7 - g u a n i d i n o h e p t a -
`2 - e n a m i d o ] - 2 - h y d r o x y e t h a n e a m i d e .
`T h e s e c o m p o u n d s have
`the s t r u c t u r e s shown
`
`in T a b l e 1 .
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 4 of 25
`
`

`
`(I) may form s a l t s w i t h
`The c o m p o u n d s of f o r m u l a
`S a l t - f o r m i n g a c i d s may be
`a c i d s .
`i n o r g a n i c or o r g a n i c
`i f
`t h e y a r e p h y s i o l o g i c a l l y a c c p e t a b l e . P r e f e r r e d
`i n o r g a n i c
`s u l f u r i c a c i d , n i t r i c a c i d
`a c i d s a r e h y d r o c h l o r i c a c i d ,
`and p h o s p h o r i c a c i d ; p r e f e r r e d o r g a n i c a c i d s
`i n c l u d e a c e -
`s u c c i n i c a c i d ,
`f u m a r i c a c i d , m a -
`t i c a c i d , p r o p i o n i c a c i d ,
`t a r t a r i c a c i d , g l u t a r i c a c i d , c i -
`l e i c a c i d , m a l i c a c i d ,
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 5 of 25
`
`

`
`t r i c
`a c i d , b e n z e n e s u l f o n i c a c i d ,
`t o l u e n s u l f o n i c
`a c i d ,
`t o l u e n e s u l f o n i c a c i d , m e t h a n e s u l f o n i c a c i d , e t h a n e -
`s u l f o n i c a c i d , p r o p a n e s u l f o n i c a c i d , a s p a r t i c a c i d a n d
`g l u t a m i c a c i d .
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 6 of 25
`
`

`
`l i s t e d a b o v e ,
`the c o m p o u n d s
`(3) a r e
`to
`(1)
`Among
`known and d e s c r i b e d
`in J a p a n e s e U n e x a m i n e d P u b l i s h e d P a t -
`(4) and
`n e t A p p l i c a t i o n No. 6 2 1 5 2 / 1 9 8 3 ; compounds
`(5) a r e
`a l s o known and d e s c r i b e d
`in J a p a n e s e U n e x a m i n e d P u b l i s h e d
`(10) and
`( 1 1 )
`P a t e n t A p p l i c a t i o n No. 4 2 3 5 6 / 1 9 8 4 ; compounds
`a r e a l s o known and d e s c r i b e d
`in J a p a n e s e U n e x a m i n e d P u b -
`l i s h e d P a t e n t A p p l i c a t i o n No. 7 6 0 4 6 / 1 9 8 4 . T h e s e known c o m -
`p o u n d s may be s y n t h e s i z e d by known m e t h o d s . C o m p o u n d s
`( 6 ) ,
`(8) and
`(9) a r e n o v e l .
`( 7 ) ,
`When
`the c o m p o u n d s of
`the p r e s e n t
`i n v e n t i o n a r e u s e d
`t h e y are a d m i n i s t e r e d e i t h e r
`i n d e -
`i m m u n o s u p p r e s s a n t s ,
`as
`in a d m i x t u r e w i t h e x c i p i e n t s or c a r r i e r s
`p e n d e n t l y or
`t o
`i n j e c t i o n s , o r a l f o r m u l a t i o n s or s u p p o s i t o r i e s .
`form
`P h a r m a c e u t i c a l l y a c c e p t a b l e e x c i p i e n t s and c a r r i e r s s h o u l d
`be s e l e c t e d and
`t h e i r
`t y p e and c o m p o s i t i o n are d e t e r m i n e d
`t h e r o u t e and m e t h o d of a d m i n i s t r a t i o n .
`I l l u s t r a t i v e
`by
`l i q u i d c a r r i e r s
`i n c l u d e w a t e r , a l c o h o l s , as w e l l as a n i -
`m a l , v e g e t a b l e and s y n t h e t i c o i l s such as s o y b e a n o i l ,
`p e a n u t o i l , s e s a m e o i l and m i n e r a l o i l s . E x e m p l a r y s o l i d
`i n c l u d e s u g a r s s u c h as m a l t o s e and s u c r o s e , a m i n o
`c a r r i e r s
`a c i d s , c e l l u l o s e d e r i v a t i v e s such as h y d r o x y p r o p y l c e l l u -
`l o s e , and o r g a n i c a c i d s a l t s s u c h as m a g n e s i u m s t e a r a t e .
`C a r r i e r s s u i t a b l e f o r u s e
`in p r e p a r i n g i n j e c t i o n s
`i n c l u d e
`p h y s i o l o g i c a l s a l i n e , b u f f e r s o l u t i o n s , s o l u t i o n s o f
`i n o s i t o l , and m a n n i t o l , a n d
`s u g a r s such as g l u c o s e ,
`g l y c o l s s u c h as e t h y l e n e g l y c o l , p r o p y l e n e g l y c o l a n d
`the c o m p o u n d s of
`t h e
`p o l y ( e t h y l e n e g l y c o l ) . A l t e r n a t i v e l y ,
`i n v e n t i o n may be f r e e z e - d r i e d
`t o g e t h e r w i t h e x c i -
`p r e s e n t
`p i e n t s s u c h as s u g a r s
`( e . g .
`i n o s i t o l , m a n n i t o l , g l u c o s e ,
`m a n n o s e , m a l t o s e and s u c r o s e ) and amino a c i d s
`( e . g . p h e n y l -
`a l a n i n e ) . B e f o r e
`such f r e e z e - d r i e d
`i n t r a v e n o u s
`i n j e c t i o n ,
`in s u i t a b l e s o l v e n t s such as s t e r i -
`c o m p o u n d s are d i s s o l v e d
`l i z e d w a t e r , p h y s i o l o g i c a l s a l i n e , g l u c o s e s o l u t i o n , e l e c -
`t r o l y t e s o l u t i o n and a m i n o a c i d s o l u t i o n . The c o n t e n t o f
`the c o m p o u n d s of
`i n v e n t i o n
`in
`the
`i m m u n o -
`the p r e s e n t
`s u p p r e s s i v e a g e n t v a r i e s w i t h
`the s p e c i f i c
`t y p e of f o r m u l a -
`i t g e n e r a l l y
`t i o n , and
`r a n g e s from 0.1
`to 100 wt%, p r e f e r -
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 7 of 25
`
`

`
`to 98 wt%. With
`t h e
`I n j e c t i o n s , t h e c o n t e n t of
`a b l y from 1
`r a n g e s from 0.1
`to 30 wt%, p r e -
`a c t i v e
`i n g r e d i e n t g e n e r a l l y
`to 10 wt%. For o r a l a d m i n i s t r a t i o n ,
`f e r a b l y from 1
`t h e c o m -
`i n v e n t i o n may be mixed w i t h any o f
`p o u n d s of
`t h e p r e s e n t
`l i s t e d a b o v e , and u s e d
`t h e
`l i q u i d c a r r i e r s
`the s o l i d or
`in
`form of
`t a b l e t s , c a p s u l e s , p o w d e r s , g r a n u l e s ,
`l i q u i d s o r
`dry s y r u p s . With c a p s u l e s ,
`t a b l e t s , g r a n u l e s and p o w d e r s ,
`the c o n t e n t of
`t h e a c t i v e
`r a n g e s g e n e r a l l y f r o m
`i n g r e d i e n t
`to 98 wt%.
`to 100 wt%, p r e f e r a b l y from 25
`5
`The d o s a g e of
`the
`t h e
`i m m u n o s u p p r e s s a n t c o n t a i n i n g
`c o m p o u n d s of
`the p r e s e n t
`i n v e n t i o n as
`the a c t i v e
`i n g r e d i e n t
`s h o u l d be p r o p e r l y d e t e r m i n e d d e p e n d i n g upon
`the age a n d
`t h e p a t i e n t , as w e l l as
`body w e i g h t of
`the s e v e r i t y a n d
`the d i s e a s e . The e f f e c t i v e dose g e n e r a l l y
`t y p e of
`r a n g e s
`to 100 m g / k g p e r day f o r p a r e n t e r a l a d m i n i s t r a t i o n ,
`from 1
`to 500 mg/kg p e r day f o r o r a l a d m i n i s t r a t i o n .
`and from 5
`The c o m p o u n d s of
`the p r e s e n t
`i n v e n t i o n a r e c h a r a c -
`low
`t o x i c i t y and s m a l l a c c u m u l a t i o n
`t e r i z e d by r e l a t i v e l y
`t o x i c i t y upon c o n t i n u o u s a d m i n i s t r a t i o n . The L D 5 0 - v a l u e s
`of
`(11)
`f o r a s i n g l e
`(1)
`of c o m p o u n d s
`to
`i n t r a p e r i t o n e a l a d -
`to m i c e ar shown
`in T a b l e 2 .
`m i n i s t r a t i o n
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 8 of 25
`
`

`
`i n v e n t i o n e x e r t
`the p r e s e n t
`The c o m p o u n d s of
`i n h i b i t o -
`t h e f u n c t i o n of
`l y m p h o c y t e s of
`ry e f f e c t s on
`i m m u n o l o g i c a l
`the m e t h o d of W a i t h e et a l .
`( W a i t h e
`i m p o r t a n c e . T e s t s by
`et a l . , H a n d b o o k of E x p e r i m e n t a l I m m u n o l o g y , page 2 6 . 1 ,
`the p r e s e n t
`r e v e a l e d
`t h e compound of
`i n v e n t i o n
`t h a t
`1 9 7 8 ) ,
`the T - l y m p h o c y t e b l a s t g e n e s i s s t i m u -
`i n h i b i t e d
`a p p r e c i a b l y
`the r e a c t i o n of B -
`l a t e d by c o n c a n a v a l i n A
`(Con A) and
`l y m p h o c y t e b l a s t g e n e s i s s t i m u l a t e d by
`l i p o p o l y s a c c h a r i d e
`( L P S ) . T h e r e f o r e
`in v i v o e x p e r i m e n t s have been c o n d u c t e d
`the e f f e c t s of
`to e x a m i n e
`the compounds of
`the p r e s e n t
`i n -
`in a c c o r d a n c e w i t h
`v e n t i o n on T - l y m p h o c y t e s . F i r s t ,
`the m e -
`t h o d of L a g r a n g e et a l .
`( L a g r a n g e et a l . , J o u r n a l of E x p e -
`r i m e n t a l M e d i c i n e , 139, 5 2 8 - 5 4 2 , 1 9 7 4 ) ,
`the c o m p o u n d s o f
`i n v e n t i o n were e x a m i n e d f o r
`t h e i r e f f e c t s o n
`the p r e s e n t
`in mice
`t h a t had b e e n s e n s i -
`d e l a y e d - t y p e h y p e r s e n s i v i t y
`red b l o o d c e l l s . The d e l a y e d - t y p e h y p e r -
`t i z e d w i t h s h e e p
`s e n s i v i t y c o u l d be b l o c k e d by a d m i n i s t e r i n g
`the c o m p o u n d
`the s e n s i t i z a t i o n w i t h
`t h r e e c o n s e c u t i v e days f o l l o w i n g
`f o r
`red b l o o d c e l l s . A d d i t i o n a l l y ,
`in a c c o r d a n c e w i t h
`t h e
`s h e e p
`m e t h o d of J e r n e et a l .
`( J e r n e et a l . , C e l l - b o u n d A n t i b o d i e s ,
`t h e c o m p o u n d s of
`the p r e s e n t
`pp. 1 0 9 - 1 2 2 , 1 9 6 3 ) ,
`i n v e n t i o n
`t h e i r e f f e c t s on
`were c h e c k e d f o r
`the p l a q u e - f o r m i n g a b i l i -
`ty of s p l e e n c e l l s from mice s e n s i t i z e d by a n t i g e n i c s h e e p
`red b l o o d c e l l s . The c o m p o u n d s of
`the p r e s e n t
`i n v e n t i o n a l -
`in
`t h i s
`so e x h i b i t e d b l o c k i n g e f f e c t s
`I t was
`t e s t .
`t h e r e -
`t h a t
`f o r e c o n f i r m e d
`t h e c o m p o u n d s of
`the p r e s e n t
`i n v e n t i o n
`e x h i b i t e d
`the a n t i b o d y p r o d u c t i o n
`i n h i b i t o r y a c t i o n on
`by B - l y m p h o c y t e s .
`The a b o v e d a t a show t h e a b i l i t y of
`the c o m p o u n d s o f
`the f u n c t i o n s of B- and T -
`i n v e n t i o n
`i n h i b i t
`to
`the p r e s e n t
`l y m p h o c y t e s . The
`i n h i b i t e d B- and T - l y m p h o c y t e s
`r e s p e c t i v e -
`t h e s u p p r e s s i o n of h u m o r a l
`i m m u n i t y and
`t h a t o f
`ly mean
`the
`c e l l - m e d i a t e d
`i m m u n i t y . T h e r e f o r e ,
`i m m u n o s u p p r e s s a n t s
`the c o m p o u n d s of
`i n v e n t i o n as
`t h e p r e s e n t
`t h e
`c o n t a i n i n g
`in s u p p r e s s i n g
`i n g r e d i e n t a r e v e r y e f f e c t i v e
`t i s s u e
`a c t i v e
`the
`t r a n s p l a n t a t i o n of an o r g a n
`t h a t a c c o m p a n i e s
`r e j e c t i o n
`or s k i n p r o b a b l y b e c a u s e of an a b n o r m a l l y e n h a n c e d h u m o r a l
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 9 of 25
`
`

`
`i m m u n i t y . The compounds can a l s o be used f o r
`or c e l l u l a r
`t r e a t m e n t of d i s e a s e s c a u s e d p r i m a r i l y by v a r i o u s f o r m s
`the
`of a u t o i m m u n i t y , as w e l l as r h e u m a t i s m and a l l e r g i c
`i n v e n t i o n seem
`d i s e a s e s . The c o m p o u n d s of
`the p r e s e n t
`t o
`t h a n
`the p r i o r a r t
`have a d i f f e r e n t m e c h a n i s m of a c t i o n
`i m -
`t h e r e f o r e no s u c h s e r i o u s s i d e e f -
`m u n o s u p p r e s s a n t s and have
`f e c t s as c a u s i n g d i s o r d e r
`i s
`in h e m a t o p o i e t i c o r g a n s as
`i m m u n o s u p p r e s s a n t s c l a s s i f i e d as c y t o t o x i c
`c a u s e d by a l l
`the d e v e l o p m e n t of g a s t r i c u l c e r s and c a t a -
`s u b s t a n c e s , or
`t h a t o f t e n a c c o m p a n i e s
`the a d m i n i s t r a t i o n of s t e r o i d
`r a c t s
`h o r m o n e s .
`r e s u l t s are p r o v i d e d f o r f u r t h e r
`t e s t
`The f o l l o w i n g
`i l l u s t r a t i o n s of
`t h e p h a r m a c o l o g i c e f f e c t s of
`the c o m p o u n d s
`i n v e n t i o n .
`the p r e s e n t
`of
`
`T e s t E x a m p l e 1
`
`I n h i b i t i o n of D e l a y e d - T y p e H y p e r s e n s i t i v i t y
`S e n s i t i z e d by Sheep Red Blood C e l l s
`
`in M i c e
`
`CDF1 mice
`f e m a l e ) w e r e s e n s i t i z e d by
`( 8 - w k - o l d ,
`i n t r a -
`i n j e c t i o n of 1 x 105 s h e e p
`red b l o o d c e l l s . F o u r
`v e n o u s
`the s e n s i t i z e d mice were s u b c u t a n e o u s l y
`l a t e r ,
`days
`i n j e c t -
`in one f o o t p a d w i t h 1 x 108 s h e e p
`red b l o o d c e l l s so a s
`ed
`i n d u c e d e l a y e d - t y p e h y p e r s e n s i t i v i t y . The s w e l l i n g o f
`to
`the 2 4 t h h o u r w i t h a m i c r o -
`the f o o t p a d was m e a s u r e d at
`the
`m e t e r . Each of
`t e s t compounds was a d m i n i s t e r e d
`i n t r a -
`in
`i n d i c a t e d
`the a m o u n t s
`in T a b l e 3
`( m g / k g )
`p e r i t o n e a l l y
`f o r a p e r i o d of 3 d a y s
`t h e day f o l l o w i n g
`t h a t s t a r t e d on
`i n i t i a l s e n s i t i z a t i o n . The d e g r e e of f o o t p a d s w e l l i n g
`t h e
`in r e l a t i v e v a l u e s , w i t h
`t h e v a l u e f o r a c o n -
`i n d i c a t e d
`was
`t r o l g i v e n p h y s i o l o g i c a l s a l i n e
`i n s t e a d of
`the
`t e s t c o m -
`t a k e n as 100%. The
`r e s u l t s a r e shown
`in T a b l e
`p o u n d s b e i n g
`the g r e a t a b i l i t y of
`from which one can see
`the c o m -
`3,
`the p r e s e n t
`p o u n d s of
`i n v e n t i o n
`to
`i n h i b i t d e l a y e d
`t y p e h y -
`p e r s e n s i t i v i t y .
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 10 of 25
`
`

`
`T e x t E x a m p l e 2
`
`I n h i b i t i o n of ConA
`g e n e s i s
`
`i n d u c e d r e a c t i o n of T - l y m p h o c y t e b l a s t
`
`S p l e e n c e l l s f r o m BALB/C mice w e r e d i s t r i b u t e d a m o n g
`t h a t each w e l l c o n t a i n e d 2 x 1 0 5
`in a m i c r o p l a t e so
`w e l l s
`c e l l s / 0 . 2 ml. Each of
`t h e s e l e c t e d
`t e s t compounds was a d d e d
`l a t t e r b e i n g u s e d as a c o n t r o l .
`to a l l w e l l s but o n e ,
`the
`(5 µ g / m l ) was a d d e d
`to a l l
`t h e w e l l s and
`ConA
`the so p r e -
`p a r e d c e l l s u s p e n s i o n s were c u l t i v a t e d
`in a 5% C02
`i n c u b a -
`t o r f o r 72 h o u r s at 3 7 ° C . E i g h t h o u r s b e f o r e
`the c o m p l e t i o n
`i n c u b a t i o n , 1 µCi of 3 H - t h y m i d i n e was added
`to e a c h
`t h e
`of
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 11 of 25
`
`

`
`the
`t h e c u l t u r e d c e l l s
`t h e u p t a k e of
`w e l l and
`t h y m i d i n e by
`l i q u i d s c i n t i l l a t i o n c o u n t e r
`was m e a s u r e d w i t h a
`to e s t i m a -
`r e a c t i o n of T - l y m p h o c y t e b l a s t g e n e s i s .
`the p r o g r e s s of
`te
`i n h i b i t i o n of
`the g e n e s i s by each of
`The p e r c e n t a g e
`t h e
`t e s t c o m p o u n d s was c a l c u l a t e d by:
`( 1 - Bdpm/Adpm) x 1 0 0
`i n d i c a t e s
`the u p t a k e c o u n t f o r
`w h e r e i n Adpm
`the a d d i t i o n
`of ConA a l o n e and Bdpm,
`the c o u n t f o r
`t h e a d d i t i o n of b o t h
`ConA and
`t e s t c o m p o u n d . The r e s u l t s a r e shown
`t h e
`in T a b l e
`4 .
`
`As T a b l e 4 s h o w s ,
`t e s t compound Nos. 1, 4, 5, 6, 8 a n d
`in a c c o r d a n c e w i t h
`11
`t h e p r e s e n t
`i n v e n t i o n e x h i b i t e d a
`the r e a c t i o n of T - l y m p h o c y t e
`to
`i n h i b i t
`s t r o n g a b i l i t y
`b l a s t g e n e s i s .
`
`T e s t E x a m p l e 3
`
`I n h i b i t i o n of LPS
`g e n e s i s
`
`i n d u c e d
`
`r e a c t i o n of B - l y m p h o c y t e b l a s t
`
`The u p t a k e of 3 H - t h y m i d i n e by B - c e l l b l a s t s was m e a s u r -
`in a c c o r d a n c e w i t h
`the m e t h o d u s e d
`in T e s t Example 2 ,
`
`ed
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 12 of 25
`
`

`
`t h a t
`r e p l a c e d by 100 ug/ml o f
`i n d u c e r ConA was
`the
`e x c e p t
`i n h i b i t i o n of b l a s t g e n e -
`LPS from E. c o l i . The p e r c e n t a g e
`s i s by s e l e c t e d
`t e s t c o m p o u n d s was a l s o d e t e r m i n e d by
`t h e
`r e s u l t s a r e
`in Text E x a m p l e 2. The
`same m e t h o d as used
`in T a b l e 5,
`from which one can see
`t h a t compound N o s .
`shown
`1, 4, 5, 6 and 11 of
`the p r e s e n t
`i n v e n t i o n were h i g h l y e f -
`the L P S - i n d u c e d g e n e s i s of B -
`f e c t i v e
`in s u p p r e s s i n g
`l y m p h o c y t e b l a s t s .
`
`T e s t E x a m p l e 4
`
`I n h i b i t i o n of P r o d u c t i o n of A n t i b o d i e s a g a i n s t S h e e p Red
`B l o o d C e l l s
`
`( 6 - 1 0 wk o l d ,
`i m m u n i z e d by
`f e m a l e ) were
`CDF1 mice
`i n -
`i n j e c t i o n of 1 x 108 s h e e p
`red b l o o d c e l l s . T h e
`t r a v e n o u s
`i n t r a p e r i t o n e a l l y a d m i n i s t e r e d 12.5 mg/kg p e r d a y
`m i c e were
`t e s t c o m p o u n d s f o r a p e r i o d of 3 days s t a r t i n g
`of s e l e c t e d
`i n t r a v e n o u s
`f r o m
`i n j e c t i o n . Four d a y s
`the day f o l l o w i n g
`the
`i s o l a t e d from
`the mice and
`t h e
`s p l e e n c e l l s were
`l a t e r ,
`n u m b e r of p l a q u e - f o r m i n g c e l l s was c o u n t e d . The p e r c e n t a g e
`i n h i b i t i o n of a n t i b o d y p r o d u c t i o n was c a l c u l a t e d b y
`the c o u n t f o r a c o n t r o l g r o u p
`( 1 - B/A) x 100 w h e r e i n A
`is
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 13 of 25
`
`

`
`f o r
`( g i v e n p h y s i o l o g i c a l s a l i n e ) and B,
`the
`t r e a t e d g r o u p .
`Compound Nos. 1, 2, 4, 5 and 6 e x h i b i t e d
`i n h i b i t i o n s o f
`9 8 . 5 % , 9 5 . 9 % , 8 9 , 4 % , 89,4% and 9 2 , 9 % ,
`r e s p e c t i v e l y .
`is c l e a r from
`As
`the f o r e g o i n g d e s c r i p t i o n ,
`the c o m -
`p o u n d s of
`the p r e s e n t
`i n v e n t i o n have a r e l a t i v e l y
`low
`t o x i -
`(LD50 b e t w e e n 1 2 . 5 and 50 m g / k g ) and e x h i b i t good
`i m -
`c i t y
`m u n o s u p p r e s s i v e a c t i o n s such as
`the
`i n h i b i t i o n of d e l a y e d -
`the
`i n h i b i t i o n of
`the f u n c t i o n s o f
`t y p e h y p e r s e n s i t i v i t y ,
`T- and B - l y m p h o c y t e s , and
`the
`i n h i b i t i o n of
`the p r o d u c t i o n
`of a n t i b o d i e s a g a i n s t s h e e p red b l o o d c e l l s . The c o m p o u n d s
`i n v e n t i o n seem
`of
`to have a d i f f e r e n t m e c h a n i s m
`t h e p r e s e n t
`of a c t i o n from
`the p r i o r a r t
`i m m u n o s u p p r e s s a n t s , and b e -
`t h i s f a c t ,
`c a u s e of
`l o w e r
`i m m u n o s u p p r e s s i n g a g e n t s h a v i n g
`s i d e e f f e c t s may be p r e p a r e d from
`the c o m p o u n d s of
`the p r e -
`i n v e n t i o n .
`s e n t
`f o l l o w i n g E x a m p l e s are p r o v i d e d f o r f u r t h e r
`The
`s t r a t i o n s of
`i n v e n t i o n .
`the p r e s e n t
`
`i l l u -
`
`E x a m p l e 1
`T h i r t y p a r t s by w e i g h t of a h y d r o c h l o r i d e of c o m p o u n d
`to make a
`No. 1 w a s mixed w i t h p u r i f i e d w a t e r
`t o t a l o f
`2 , 0 0 0 p a r t s . The s o l u t i o n was p a s s e d
`t h r o u g h a M i l l i p o r e
`f i l t e r of GS
`t y p e f o r s t e r i l i z a t i o n p u r p o s e s . Two grams o f
`i n t o 10-ml v i a l s and f r e e z e - d r i e d
`the f i l t r a t e was put
`t o
`i n j e c t i o n s e a c h c o n t a i n i n g 30 mg of
`the h y d r o c h l o -
`p r e p a r e
`(1) p e r v i a l .
`r i d e of compound
`
`E x a m p l e 2: G r a n u l e s
`i n t i m a t e m i x t u r e of 50 p a r t s by w e i g t h of a h y d r o -
`An
`( 2 ) , 600 p a r t s of
`c h l o r i d e of compound
`l a c t o s e , 330 p a r t s
`of c r y s t a l l i n e c e l l u l o s e and 20 p a r t s of h y d r o x y p r o p y l c e l -
`( R )
` a n d
`l u l o s e was c o m p a c t e d w i t h a R o l l e r C o m p a c t o r
`i n t o p a r t i c l e s w h i c h were s i e v e d
`to p r o v i d e g r a n u l e s
`g r o u n d
`of a s i z e b e t w e e n 16 and 60 m e s h .
`
`E x a m p l e 3: T a b l e t s
`A m i x t u r e of 30 p a r t s by w e i g h t of a h y d r o c h l o r i d e o f
`
`NOVARTIS EXHIBIT 2029
`Par v Novartis, IPR 2016-00084
`Page 14 of 25
`
`

`
`( 3 ) , 120 p a r t s of c r y s t a l l i n e
`l a c t o s e , 147 p a r t s
`compound
`of c r y s t a l l i n e c e l l u l o s e and 3 p a r t s of m a g n e s i u m s t e a r a t e
`was p r o c e s s e d w i t h a V - t y p e p e l l e t i z i n g m a c h i n e
`to p r o d u c e
`t a b l e t s e a c h w e i g h i n g 300 mg.
`The
`f o l l o w i n g R e f e r e n c e E x a m p l e s are g i v e n
`to show
`t h e
`the n o v e l c o m p o u n d s
`m e t h o d s of s y n t h e s i z i n g
`in a c c o r d a n c e
`the R e f e r e n c e E x a m p l e s , d i Z ,
`i n v e n t i o n .
`the p r e s e n t
`In
`w i t h
`TAD, and GP
`t r i a z a d e c a n e a n d
`r e p r e s e n t d i b e n z y l o x y c a r b o n y l ,
`g u a n i d