Covalent analogues of DNA base-pairs and triplets IV+. Synthesis of trisubstituted benzenes bearing purine and[s]or pyrimidine rings by cyclotrimerization of 6-ethynylpurines and[s]or 5-ethynyl-1,3-dimethyluracil

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DNA Base-Pairs and Triplets

1223

COVALENT ANALOGUES OF DNA BASE-PAIRS AND TRIPLETS IV+. SYNTHESIS OF TRISUBSTITUTED BENZENES BEARING PURINE AND/ OR PYRIMIDINE RINGS BY CYCLOTRIMERIZATION OF 6 -ETHYNYLPURINES AND/ OR 5 -ETHYNYL-1 ,3 -DIMETHYLURACIL Michal HOCEKa1,*, Irena G. STARÁa2, Ivo STARÝa3 and Hana DVOŘÁKOVÁb a

b

Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, CZ-16610 Prague 6, Czech Republic; e-mail: 1 [email protected], 2 [email protected], 3 [email protected] Central Laboratory of NMR, Institute of Chemical Technology, Prague, CZ-16628 Prague 6, Czech Republic; e-mail: [email protected]

Received May 20, 2002 Accep t ed Ju n e 16, 2002

Ni-Cat alyzed cyclo t rim erizat io n s o f 6-et h yn ylp u rin es 3 o r 5-et h yn yl-1,3-d im et h ylu racil (4 ) affo rd ed t h e 1,2,4-t ris(p u rin -6-yl)b en zen es 7 o r 1,2,4-t ris(1,3-d im et yh ylu racil-5-yl)b en zen e (9 ), resp ect ively. Th e sym m et rical 1,3,5-t ris(p u rin -6-yl)ben zen es 8 were also fo rm ed as m in o r p ro d u ct s in very lo w yield s. Co -cyclo t rim erizat io n o f 9-ben zyl-6-et h yn ylp u rin e (3 a) wit h 4 affo rd ed t h e t ris(p u rin yl)b en zen e 7 a as a m ajo r p ro d u ct alo n g wit h 1,2-b is(9-b en zylp u rin 6-yl)-4-(1,3-d im et h ylu racil-5-yl)b en zen e (1 0 ) an d a co m p lex m ixt u re o f o t h er d erivat ives an d iso m ers. Co m p o u n d s 7 –1 0 are an alo gu es o f Ho o gst een base-t rip let s. Ke y w o rd s: Pu rin es; Pyrim id in es; N u cleo b a ses; H o o gst een t rip let s; C yclo t rim eriza t io n s; Nickel; Alkyn es; [2+2+2] Cyclo ad d it io n s.

Two m ain h yd ro gen bo n d in g m o t ifs exist in DNA: t h e W at so n –Crick m o t if in d u p lexes an d t h e Ho o gst een m o t if in t rip lexes. Hyd ro gen bo n d in g is cru cial t o t h e abilit y o f t h e t wo st ran d s t o st ay an n ealed t o each o t h er bu t eq u ally im p o rt an t is t h e abilit y t o sep arat e fro m o n e an o t h er in t h e righ t m o m en t . Th erefo re t h e effect o f m an y clin ically u sed an t it u m o r agen t s is b ased o n DNA cro ss-lin kin g 1 o r o n in t ercalat io n 2 in t o DNA. Nu m ero u s m o d els an d an alo gu es o f W at so n –Crick base p airs co n sist in g o f an n elat ed 3 o r cro ss-lin ked 4 p u rin e an d p yrim id in e h et ero cycles o r even m o re sim p le aro m at ic rin gs5,6 h ave been p rep ared . Su ch base-p air an alo gu es m ay in t eract wit h DNA (e.g. by in t ercalat io n ); if in co rp o rat ed in t o sin gle-st ran d ed + Fo r Part III see ref. 9

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DNA, t h ey are co m p lem en t ary t o abasic sit e o f a d am aged DNA st ran d ; o r alt ern at ively, if in co rp o rat ed t o a d u p lex, t h ey fo rm p erm an en t cro ss-lin ks. O n t h e o t h er h an d , n o syst em at ic research o n co valen t an alo gu es o f an o t h er im p o rt an t DNA H-b o n d in g m o t if, Ho o gst een t rip let s (Fig. 1), h as been rep o rt ed so far, t rip u rin ylam in es bein g t h e o n ly kn o wn exam p le 7 . Very recen t ly, we h ave p rep ared a n ew t yp e o f co valen t base-p air an alo gu es co n sist in g o f vario u s p u rin e d im ers an d p u rin e-p yrim id in e co n ju gat es lin ked t h ro u gh p o sit io n s 6 an d 6′ (o r p o sit io n 5 o f p yrim id in e) by acet ylen e, d iacet ylen e, vin ylen e an d et h ylen e 8 as well as para- o r m etap h en ylen e 9 lin kers. Su ch carbo n lin kers co n n ect ed t o carbo n at o m s o f t h e h et ero cycles were exp ect ed t o be st able t o ward s en zym at ic d egrad at io n . Sign ifican t cyt o st at ic act ivit y h as b een fo u n d 8 in so m e b is(p u rin -6-yl)acet ylen es an d d iacet ylen es, wh ile t h e p art ially an d fu lly sat u rat ed d erivat ives, p h en ylen e-lin ked an alo gu es as well as t h e p u rin e-p yrim id in e co n ju gat es were in act ive. Takin g also in t o acco u n t t h e h igh cyt o st at ic act ivit y o f Natural nucleobase-triplets (examples): R N CH3 N R N N H O N O N N H H H H N N H N N H O O CH3 N N N H R N H N R N N N N O O R T.AT triplet A.AT triplet

CH3

N R

Covalent triplet analogues:

R N

O

O

N

N

N

N

N

N

N

N

N

O

R N

N N

O

O

N N

N

R N

N

O

N

N

O

N

N N

O

N

N N

N

N R

R

FIG . 1 St ru ct u re o f n at u ral Ho o gst een t rip let s an d t h eir co valen t an alo gu es u n d er st u d y

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6-arylp u rin e ribo n u cleo sid es10 , we h ave d ecid ed t o p rep are a n ew t yp e o f Ho o gst een t rip let s an alo gu es co n sist in g o f b en zen e rin gs b earin g t h ree n u cleo bases (p u rin es o r p yrim id in es). A p relim in ary co m m u n icat io n o n t h e syn t h esis o f 1,3,4- an d 1,3,5-t ris(p u rin -6-yl)ben zen es h as recen t ly been p u blish ed 11 . Th e p resen t fu ll-p ap er gives t h e resu lt s in fu ll d et ails an d ext en d s t h e st u d y t o ward s t h e an alo gu es bearin g p u rin e an d / o r p yrim id in e rin gs (Fig. 1). R'

H for R' = SiMe3

Cl N

N

i)

N

N R 2a-2c

N

N

N R

1a-1e

R

a

a

b

Bn SiMe3 THP SiMe3

b

Bn THP

c

Me

SiMe3

c

Me

d

Bn

Bu

e

Bn

Ph

2,3

ii)

N

N N 3a-3c

N R

R

1

R'

N

i) R'C≡CH, Pd(PPh3)4, CuI, Et3N, DMF, 120 °C, 7 h; ii) NH3/MeOH, r.t., 3 h

SCHEME 1

O u r o rigin al ap p ro ach 11 t o t h e t rip let an alo gu es relies o n cyclo t rim erizat io n o f p ro p erly fu n ct io n alized alkyn es. Su bst it u t ed 6-et h yn ylp u rin es 1 are read ily available by t h e So n o gash ira co u p lin g o f 6-ch lo ro p u rin e d erivat ives 2 wit h alkyn es (an alo gy t o t h e kn o wn 1 2 p ro ced u re fo r 6-alkyn yl9-p h en ylp u rin e d erivat ives). Th e t erm in al acet ylen es 3 were p rep ared in go o d yield s in t wo st ep s co n sist in g in t h e co u p lin g o f 2 wit h (t rim et h ylsilyl)acet ylen e fo llo w ed b y d esilylat io n u sin g m et h an o lic am m o n ia 1 2 (Sch em e 1 ). An alo go u s ap p ro ach h as b een ap p lied t o t h e syn t h esis o f 5-et h yn yl-1,3-d im et h ylu racil (4 ) st art in g fro m 5-io d o -1,3-d im et h ylu racil (5 ) (Sch em e 2). W h ile t h e co u p lin g o f 5 wit h (t rim et h ylsilyl)acet ylen e t o give t h e kn o wn 13 TMS-p ro t ect ed acet ylen e 6 p ro ceed ed q u it e well, su bseq u en t d esilylat io n o f 6 u sin g m et h an o lic am m o n ia d id n o t lead t o t h e t erm in al acet ylen e 4 bu t t o a co m p lex m ixt u re o f o ligo - an d / o r p o lym ers. Eq u ally u n su ccessfu l were at t em p t s u sin g KF/ m et h an o l o r K2 CO 3 / m et h an o l. Fin ally we h ave su cceed ed m akin g u se o f TBAF in THF wh ich gave t h e d esired acet ylen e 4 in a m o d erat e yield o f 51%.

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O

O I

N O

i)

5

ii)

N O

N

O

SiMe3 N O

N 6

H

N 4

i) Me3SiC≡CH, Pd(PPh3)4, CuI, Et3N, DMF, 100 °C, 8h; ii) TBAF,THF, 0 °C then r.t., 5 h

SCHEME 2

Th e 9-su b st it u t ed 6-et h yn ylp u rin es 3 a an d / o r 3 b were u sed as m o d el su bst rat es fo r a series o f cyclo t rim erizat io n exp erim en t s varyin g t ran sit io n m et al cat alyst s an d react io n co n d it io n s acco rd in g t o lit erat u re p ro t oco ls 1 4 –1 9 (Sch em e 3 ). W h ile TaC l 5 in b en zen e, t h e G ru b b s cat alyst Ph CH=Ru (PCy 3 )Cl 2 in d ich lo ro m et h an e, Pd (PPh 3 ) 4 o r Ni(CO ) 2 (PPh 3 ) 2 in t et rah yd ro fu ran , an d t h e W ilkin so n cat alyst Rh Cl(PPh 3 )3 in et h an o l left t h e st art in g alkyn es 3 a an d / o r 3 b u n t o u ch ed even u n d er reflu x fo r a p ro lo n ged react io n p erio d , t h e u se o f Cp Co (CO )2 in d ecan e at 140 °C wit h t h e co n co m it an t visible ligh t irrad iat io n led t o a very co m p lex m ixt u re co n t ain in g o n ly t races o f t h e t arget p ro d u ct s 7 o r 8 (acco rd in g t o t h e MS an alysis o f t h e cru d e react io n m ixt u re). Th e o bserved lo w react ivit y o f t h ese alkyn es t o ward s cyclo t rim erizat io n m igh t be exp lain ed in t erm s o f a su bst an t ial d ecrease o f t h e elect ro n d en sit y at t h e t rip le bo n d d u e t o t h e p resen ce o f t h e elect ro n -d eficien t p u rin e m o iet y. Th e sit u at io n d ram at ically ch an ged wh en ap p lyin g a h igh ly react ive Ni(CO D) 2 (CO D = cyclo o ct a-1,4-d ien e) co m p lex t o en fo rce t h e t rim erizat io n (Sch em e 3, Table I). Th e THP-p ro t ect ed 3 b wit h a cat alyt ic am o u n t o f Ni(CO D)2 an d PPh 3 affo rd ed an 8 : 1 m ixt u re o f t ris(p u rin -9-yl)ben zen es 7 b an d 8 b in go o d yield (74%; Table I, En t ry 2). Bo t h regio iso m ers were su ccessfu lly sep arat ed by co lu m n ch ro m at o grap h y. Th e u se o f a st o ich io m et ric am o u n t o f Ni(CO D)2 (1/ 3 eq u ivalen t ) wit h o u t PPh 3 as a st abilizin g ligan d gave a 4 : 1 m ixt u re o f 7 b an d 8 b in m o d erat e yield (41%; Table I, En t ry 3). Co m p o u n d 7 b was d ep ro t ect ed by m ean s o f wet Do wex 50X8 (H + fo rm ) in m et h an o l 20 t o give t h e free p u rin e d erivat ive 7 f in 70% yield . An alo go u sly, t h e react io n o f t h e Bn -p ro t ect ed co m p o u n d 3 a w it h Ni(CO D)2 an d PPh 3 affo rd ed t h e u n sym m et rical 1,2,4-t ris(p u rin -9-yl)ben zen e 7 a in 50% yield , wh ile t h e sym m et rical p ro d u ct 8 a co u ld n o t b e iso lat ed in a p u re fo rm (Tab le I, En t ry 1 ). Sim ilarly, t h e react io n o f 6-et h yn yl-9-m et h ylp u rin e (3 c) gave t h e u n sym m et rical t rim er 7 c in 43% yield , wh ile t h e sym m et rical t rim er 8 c was ju st d et ect ed in sp o t s an d co u ld n o t be iso lat ed (Table I, En t ry 4).

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N H

NiL2, ligand

N

3

N

N

N

N

N

N

R N

N N

N +

N R

N

N

R N

N

N

N

R

R N

N

N

R

N 7

b

Bn THP

c

Me

7f

H

3, 7, 8 a

N

N

N N R

N R

8

Dowex 50 (H+)

SCHEME 3 TABLE I Cyclo t rim erizat io n s o f 6-et h yn ylp u rin es 1 an d 3 Yield , % En t ry

1

St art in g co m p o u n d

3a

Cat alyst , ligan d (m o le %)

Ni(CO D)2 (20), PPh 3 (50)

7

8

50

5a

2

3b

Ni(CO D)2 (20), PPh 3 (50)

66

8

3

3b

Ni(CO D)2 (33)

33

8

4

3c

Ni(CO D)2 (20), PPh 3 (50)

43

t races

5

1d

Ni(CO D)2 (20), PPh 3 (50)

o n ly t races o f cyclo t rim ers

6

1e

Ni(CO D)2 (20), PPh 3 (50)

o n ly t races o f cyclo t rim ers

7

3a

NiCp 2 (20), PPh 3 (50)

35

t races

a Co m p o u n d 8 a was n o t iso lat ed in a p u re fo rm (yield est im at ed fro m 1 H NMR o f t h e cru d e react io n m ixt u re).

Th e d isu b st it u t ed acet ylen es 1 d an d 1 e w ere also su b ject ed t o t h e cyclo t rim erizat io n u sin g cat alyt ic am o u n t o f Ni(CO D)2 an d PPh 3 . Ho wever, t h e react io n was very slu ggish t o fo rm t h e t rim ers in t race am o u n t s o n ly (MS d et ect io n ) even aft er p ro lo n ged react io n t im es an d / o r at elevat ed t em p erat u re (u p t o 60 °C). An alo go u sly t o t h e p u rin es, t h e 5-et h yn yl-1,3-d im et h ylu racil (4 ) was also cyclo t rim erized u sin g Ni(CO D)2 t o give t h e u n sym m et rical t rim er 9 in 25%

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yield (Sch em e 4). Th is react io n was m u ch slo wer t h an t h o se o f p u rin e d erivat ives, p ro bably d u e t o lo w so lu bilit y o f t h e st art in g co m p o u n d 4 in THF. Fu rt h erm o re, we h ave also t ried a co -cyclo t rim erizat io n o f p u rin e 3 a wit h p yrim id in e 4 in o rd er t o get m ixed p u rin e-p yrim id in e co n ju gat es lin ked by a ben zen e rin g. Th e co -cyclo t rim erizat io n o f 3 a an d 4 in t h e 1 : 1 rat io gave t h e u n sym m et rical p u rin e h o m o -t rim er 7 a as m ajo r p ro d u ct (25% yield ), acco m p an ied by t h e u n react ed 4 (30%) an d a co m p lex m ixt u re o f h o m o an d h et ero t risu b st it u t ed b en zen es. O u t o f t h is m ix t u re, o n ly 1 ,2 -b is(9-b en zylp u rin -6-yl)-4-(1,3-d im et h ylu racil-5-yl)b en zen e (1 0 ) was su ccessfu lly iso lat ed in p u re fo rm in 5% yield . Th is resu lt was n o t su rp rizin g wh en t akin g in t o acco u n t t h e lo wer so lu bilit y an d / o r react ivit y o f 4 . Nevert h eless, co m p o u n d 1 0 co u ld be co n sid ered as t h e first an alo gu e o f t h e real Pu ·Py·Pu Ho o gst een t rip let . O

H Ni(COD)2, PPh3

N H

O

O

N

N

O

N N 3a

Bn N

N

O

N 4

N

O

N

9

N

Ni(COD)2, PPh3

N Bn

N

N

N Bn

N

O

O

Bn N

N N

N

N N

N N

O

N

+

N

N N

O

N N N Bn

N

N

7a (25%)

+

N Bn

complex mixture of other derivatives and isomers

10 (5%)

SCHEME 4

Recen t ly, t h e u se o f st able n ickelo cen e (NiCp 2 ) in st ead o f ext rem ely airsen sit ive Ni(CO D) 2 in co u p lin g an d cyclizat io n react io n s h as b een d escribed 21 . In an alo gy, we h ave u sed NiCp 2 / PPh 3 cat alyt ic syst em fo r t h e cyclo t rim erizat io n o f 3 a t o o bt ain t h e u n sym m et rical t rim er 7 a in 35% yield (Table I, En t ry 7). Th o u gh t h e yield was so m ewh at lo wer, d u e t o m u ch easier h an d lin g o f N iC p 2 in co m p ariso n t o N i(C O D) 2 , t h is alt ern at ive m et h o d co u ld b e also ad van t ageo u sly u sed fo r t h e t rim erizat io n o f 6-et h yn ylp u rin es.

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W h ile t h e NMR sp ect ra o f t h e 1,3,5-t risu bst it u t ed ben zen e 8 b d isp layed very sim p le p at t ern s d u e t o t h eir h igh sym m et ry, t h e sp ect ra o f t h e 1,2,4-t risu bst it u t ed d erivat ives 7 a–7 c an d 7 f co n t ain ed d ist in ct set s o f sign als belo n gin g t o each p u rin e rin g. Po ssessin g ch iralit y cen t ers at t h e THP p ro t ect in g gro u p s, co m p o u n d s 7 b an d 8 b h ave t o o ccu r as m ixt u res o f d iast ereo iso m ers. In sp it e o f t h is, t h ese m at erials were ch ro m at o grap h ically h o m o gen eo u s. Alt h o u gh in t h e 1 H an d 13 C NMR sp ect ra o f 7 b so m e sign als o f t h e p ro xim al p u rin es were sp lit , t h e sp ect ra o f 8 b exh ibit ed a p erfect sym m et ry. Fu rt h erm o re, n o h in d ered ro t at io n was o bserved in d yn am ic NMR exp erim en t s wit h co m p o u n d s 7 a, 7 b an d 8 b even at lo w t em p erat u res (d o wn t o –70 °C) in d icat in g t h at t h ese co m p o u n d s co u ld easily ad o p t a p lan ar co n fo rm at io n wh ich is n ecessary fo r in t ercallat io n in t o DNA. In co n clu sio n , t h e Ni-cat alyzed cyclo t rim erizat io n s o f 6-et h yn ylp u rin es 3 p ro vid ed t h e u n sym m et rical (m ajo r) an d sym m et rical (m in o r) t ri(p u rin 6 -yl)b en zen es 7 an d 8 as t h e n o vel H o o gst een -t rip let an alo gu es. Th is m et h o d is esp ecially su it ab le fo r t h e syn t h esis o f 1,2,4-t ris(p u rin -6-yl)ben zen es fro m t erm in al et h yn ylp u rin es. An alo go u s cyclo t rim erizat io n o f 5-et h yn yl-1,3-d im et h ylu racil (4 ) gave also t h e u n sym m et rical t rim er 9 . Th e co -t rim erizat io n o f 3 a an d 4 led t o a co m p lex m ixt u re co n t ain in g t h e u n sym m et rical p u rin e h o m o -t rim er 7 a as a m ajo r p ro d u ct an d , t h erefo re, it is n o t ap p licable fo r t h e p rep arat ive syn t h esis o f m ixed t rim ers co n t ain in g bo t h p u rin e an d p yrim id in e su bst it u en t s. Th e t arget t rip let -an alo gu es 7 –1 0 were t est ed fo r t h eir cyt o st at ic act ivit y (in h ibit io n o f cell gro wt h o f t h e fo llo win g cell cu lt u res: m o u se leu kem ia L1210 cells (ATCC CCL 219), m u rin e L929 cells (ATCC CCL 1), h um an cervix carcin om a HeLaS3 cells (ATCC CCL 2.2) an d h u m an T lym p h oblastoid CCRF-CEM cell lin e (ATCC CCL 119)). Non e of th e com pou n ds exh ibited an y con siderable activity in th ese assays22 . EXPERIMENTAL Un less st at ed o t h erwise, so lven t s were evap o rat ed at 40 °C/ 2 kPa an d co m p o u n d s were d ried at 60 °C/ 2 kPa o ver P 2 O 5 . Melt in g p o in t s were d et erm in ed o n a Ko fler b lo ck an d are u n co rrect ed . NMR sp ectra were m easu red on a Bru ker AMX-3 400 (400 MHz for 1 H an d 100.6 MHz fo r 13 C), a Bru ker DRX 500 (500 MHz fo r 1 H an d 125.8 MHz fo r 13 C). Ch em ical sh ift s (δ) are given in p p m , co u p lin g co n st an t s (J) in Hz. TMS was u sed as in t ern al st an d ard . Mass sp ect ra were m easu red o n a ZAB-EQ (VG An alyt ical) sp ect ro m et er u sin g FAB (io n izat io n b y Xe, accelerat in g vo lt age 8 kV, glycero l m at rix). To lu en e was d egassed in vacuo an d st o red o ver m o lecu lar sieves u n d er argo n . DMF was d ist illed fro m P 2 O 5 , d egassed in vacuo an d st o red o ver

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m o lecu lar sieves u n d er argo n . THF was reflu xed wit h Na an d ben zo p h en o n e u n d er argo n an d fresh ly d ist illed p rio r t o u se. So n o gash ira React io n s o f 6-Ch lo ro p u rin es wit h (Trim et h ylsilyl)acet ylen e. Gen eral Pro ced u re DMF (10 m l) an d Et 3 N (4 m l) were ad d ed t h ro u gh sep t u m t o an argo n p u rged flask co n t ain in g a 6-ch lo ro p u rin e 2 (6 m m o l), TMSC≡CH (980 m g, 10 m m o l), Cu I (100 m g, 0.5 m m o l) an d Pd (PPh 3 ) 4 (100 m g, 0.087 m m o l). Th e m ixt u re was t h en st irred at 120 °C fo r 7 h an d left at am bien t tem peratu re overn igh t. Th e solven ts were evap orated in vacuo an d th e p rodu cts isolated by colu m n ch rom atograph y on silica gel (150 g, eth yl acetate/ ligh t p etroleu m 1 : 2). 9-Benzyl-6-[(trim ethylsilyl)ethynyl]purine (1 a). W h it e cryst als, yield 70%; m .p . 126–128 °C (h ep t an e/ CH 2 Cl 2 ). EI MS, m /z (rel.%): 306 (60) [M], 291 (42) [M – Me], 91 (100). 1 H NMR (400 MHz, CDCl 3 ): 0.33 (s, 9 H, (CH 3 ) 3 Si); 5.44 (s, 2 H, CH 2 ); 7.27–7.36 (m , 5 H, H-aro m .); 8.09 (s, 1 H, H-8); 8.95 (s, 1 H, H-2). 13 C NMR (100 MHz, CDCl 3 ): –0.44 ((CH 3 ) 3 Si); 47.38 (CH 2 ); 98.47, 105.46 (C≡C); 127.74, 128.68, 129.16 (CH-aro m .); 134.22 (C-aro m .); 134.84 (C-5); 141.30 (C-6); 145.21 (C-8); 151.82 (C-4); 152.70 (C-2). Fo r C 17 H 18 N 4 (306.4) calcu lat ed : 66.63% C, 5.92% H, 18.28% N; fo u n d : 66.48% C, 6.01% H, 18.26% N. 9 -(T etrah ydropyran -2 -yl)-6 -[(trim eth ylsilyl)eth yn yl]purin e (1 b ). W h it e cryst als, yield 7 8 % ; m .p . 129–131 °C (h ep t an e/ CH 2 Cl 2 ). EI MS, m /z (rel.%): 300 (37) [M], 272 (20), 217 (100) [M + H – THP], 201 (63), 85 (95) [THP]. 1 H NMR (500 MHz, CDCl 3 ): 0.34 (s, 9 H, (CH 3 ) 3 Si); 1.75–2.17 (m , 6 H, CH 2 ); 3.79 (d t , 1 H, J = 2.5, 11.7, H-5′a); 4.19 (m , 1 H, H-5′b ); 5.80 (d d , 1 H, J = 10.4, 2.4, H-1′); 8.34 (s, 1 H, H-8); 8.92 (s, 1 H, H-2). 13 C NMR (75 MHz, CDCl 3 ): 0.225 ((CH 3 ) 3 Si); 23.31, 25.44, 32.46 (CH 2 ); 69.47 (CH 2 -5′); 82.77 (CH-1′); 99.07, 106.08 (C≡C); 135.02 (C-5); 141.88 (C-6); 143.88 (C-8); 151.58 (C-4); 153.11 (C-2). Fo r C 15 H 20 N 4 O Si (300.4) calcu lat ed : 59.97% C, 6.71% H, 18.65% N; fo u n d : 59.61% C, 6.74% H, 18.41% N. 9-Methyl-6-[(trim ethylsilyl)ethynyl]purine (1c). Brown ish crystals, yield 75%; m .p . 155–158 °C (h ep t an e/ CH 2 Cl 2 ). EI MS, m /z (rel.%): 230 (42) [M], 215 (100). IR (KBr): ν = 2 960, 2 157, 1 581, 1 505, 1 443, 1 394, 1 326 cm –1 . 1 H NMR (400 MHz, CDCl 3 ): 0.32 (s, 9 H, (CH 3 ) 3 Si); 3.90 (s, 3 H, CH 3 ); 8.08 (s, 1 H, H-8); 8.91 (s, 1 H, H-2). 13 C NMR (100.6 MHz, CDCl 3 ): –0.45 ((CH 3 ) 3 Si); 29.84 (CH 3 ); 98.45, 105.31 (C≡C); 134.15 (C-5); 141.10 (C-6); 145.86 (CH-8); 152.06 (C-4); 152.52 (CH-2). For C 11 H 14 N 4 Si (230.3) calcu lated: 57.36% C, 6.13% H, 24.32% N; fo u n d : 57.04% C, 6.16% H, 24.23% N. 9-Benzyl-6-(phenylethynyl)purine23 (1 d ). Bro wn ish o il t h at so lid ified t o cryst als o n d ryin g, yield 71%; m .p . 114–117 °C. EI MS, m /z (rel.%): 310 (73) [M], 91 (89), 57 (100), 43 (86). 1 H NMR (400 MHz, CDCl 3 ): 5.48 (s, 2 H, CH 2 Ph ); 7.26–7.75 (m , 10 H, H-aro m .); 8.13 (s, 1 H, H-8); 9.00 (s, 1 H, H-2). 13 C NMR (100 MHz, CDCl 3 ): 47.41 (CH 2 Ph ); 84.13, 98.41 (C≡C); 121.38 (C-aro m .); 127.81, 128.39, 128.70, 129.19, 129.88, 132.66 (CH-aro m .); ca 134, 134.83 (C-5 an d C-aro m .); 141.90, 151.66 (C-4 an d C-6); 145.00 (CH-8); 152.77 (CH-2). EI HRMS, fo u n d : 310.1215; C 20 H 14 N 4 [M] req u ires: 310.1218. Fo r C 20 H 14 N 4 (310.4) calcu lat ed : 77.40% C, 4.55% H, 18.05% N; fo u n d : 77.23% C, 4.68% H, 17.70% N. 9-Benz yl-6-(hex-1-yn-1-yl)purine (1 e). Bro wn ish o il t h at so lid ified t o cryst als o n d ryin g, yield 77%; m .p . 71–73 °C. EI MS, m /z (rel.%): 290 (40) [M], 261 (20), 248 (56), 199 (22), 91 (100). 1 H NMR (400 MHz, CDCl 3 ): 0.95 (t , 3 H, J = 7.3, CH 3 ); 1.48–1.56 (m , 2 H, CH 2 ); 1.65–1.73 (m , 2 H, CH 2 ); 2.60 (t , 2 H, J = 7.1, CH 2 C≡); 5.44 (s, 2 H, CH 2 Ph ); 7.26–7.37 (m , 5 H, H-aro m .); 8.06 (s, 1 H, H-8); 8.93 (s, 1 H, H-2). 13 C NMR (100 MHz, CDCl 3 ): 13.56 (C H 3 ); 1 9 .6 1 , 2 2 .1 0 , 3 0 .1 3 (C H 2 ); 4 7 .3 4 (C H 2 Ph ); 7 6 .0 5 , 1 0 1 .6 5 (C ≡C ); 1 2 7 .7 9 , 1 2 8 .6 6 ,

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129.16 (CH-aro m .); 134.19, 134.90 (C-5 an d C-aro m .); 142.54, 151.45 (C-4 an d C-6); 144.73 (CH-8); 152.75 (CH-2). EI HRMS, fo u n d : 290.1512; C 18 H 18 N 4 [M] req u ires: 290.1531. Fo r C 18 H 18 N 4 (290.4) calcu lat ed : 76.46% C, 6.25% H, 19.30% N; fo u n d : 74.19% C, 6.32% H, 18.92% N. Desilylat io n o f 6-[(Trim et h ylsilyl)et h yn yl]p u rin es. Gen eral Pro ced u re A TMS d erivat ive 1 a–1 c (10 m m o l) was t reat ed wit h sat u rat ed et h an o lic am m o n ia (100 m l) fo r 3 h , t h e so lven t was evap o rat ed an d t h e p ro d u ct s were iso lat ed b y co lu m n ch ro m at o grap h y o n silica gel (150 g, et h yl acet at e). 9 -Ben z yl-6 -eth yn ylpurin e (3 a). W h it e cryst a ls, yield 6 5 % ; m .p . 1 5 8 –1 6 0 °C (h ep t a n e/ CH 2 Cl 2 ). EI MS, m /z (rel.%): 234 (84) [M]; 91 (100). 1 H NMR (500 MHz, CDCl 3 ): 3.72 (s, 1 H, ≡CH); 5.46 (s, 2 H, CH 2 ); 7.30–7.37 (m , 5 H, H-aro m .); 8.12 (s 1 H, H-8); 8.99 (s, 1 H, H-2). 13 C NMR (125 MHz, CDCl 3 ): 47.47 (CH 2 ); 77.94 (C≡); 86.08 (≡CH); 127.89, 128.78, 129.23 (C H -a ro m .); 1 3 4 .7 0 (C -5 ); 1 4 0 .6 8 (C -6 ); 1 4 5 .4 6 (C -8 ); 1 5 1 .7 5 (C -4 ); 1 5 2 .7 3 (C -2 ). Fo r C 14 H 10 N 4 (234.2) calcu lat ed : 71.78% C, 4.30% H, 23.92% N; fo u n d : 71.46% C, 4.37% H, 23.79% N. 6-Ethynyl-9-(tetrahydropyran-2-yl)purine (3 b). W h it e cryst als, yield 70%; m .p . 105–108 °C (h ep t an e/ CH 2 Cl 2 ). EI MS, m /z (rel.%): 228 (26) [M], 200 (18) [M + H – THP], 145 (48), 85 (100) [THP]. 1 H NMR (400 MHz, CDCl 3 ): 1.69–2.19 (m , 6 H, CH 2 ); 3.72 (s, 1 H, ≡CH); 3.80 (d t , 1 H, J = 2.6, 11.5, H-5′a); 4.16–4.22 (m , 1 H, H-5′b); 5.81 (d d , 1 H, J = 10.2, 2.4, H-1′); 8.36 (s, 1 H, H-8); 8.95 (s, 1 H, H-2). 13 C NMR (75 MHz, CDCl 3 ): 23.31, 25.44, 32.46 (CH 2 ); 69.51 (CH 2 -5′); 78.59 (≡C); 82.83 (CH-1′); 86.75 (≡CH); 135.67 (C-5); 141.30 (C-6); 144.19 (C-8); 151.55 (C-4); 153.16 (C-2). Fo r C 12 H 12 N 4 O (228.3) calcu lat ed : 63.15% C, 5.30% H, 24.55% N; fo u n d : 63.19% C, 5.01% H, 24.26% N. 6 -Eth yn yl-9 -m eth ylpurin e (3 c ). W h it e cryst a ls, yield 7 7 % ; m .p . 2 2 0 –2 2 2 °C (h ep t a n e/ CH 2 Cl 2 ). EI MS, m /z (rel.%): 158 (100) [M]. IR (KBr): ν = 3 155, 3 097, 2 101, 1 579, 1 504, 1 437, 1 425, 1 345 cm –1 . 1 H NMR (400 MHz, CDCl 3 ): 3.71 (s, 1 H, ≡CH); 3.94 (s, 3 H, CH 3 ); 8.13 (s, 1 H, H-8); 8.97 (s, 1 H, H-2). 13 C NMR (100.6 MHz, CDCl 3 ): 29.87 (CH 3 ); 77.95 (C≡); 85.96 (≡CH); 134.78 (C-5); 140.55 (C-6); 146.11 (CH-8); 152.03 (C-4); 152.57 (CH-2). Fo r C 8 H 6 N 4 (158.1) calcu lat ed : 60.75% C, 3.82% H, 35.42% N; fo u n d : 60.49% C, 3.79% H, 35.09% N. 1,3-Dim et h yl-5-[(t rim et h ylsilyl)et h yn yl]p yrim id in e-2,4-(1H,3H)-d io n e (6 ) DMF (9 m l) an d Et 3 N (3 m l) were ad d ed t o an argo n p u rged flask co n t ain in g 5 (2.66 g, 10 m m o l), TMSC≡CH (2.1 g, 21 m m o l), Cu I (160 m g, 5.2 m m o l) an d Pd (PPh 3 ) 4 (160 m g, 0.14 m m o l). Th e m ixt u re was st irred u n d er argo n at 100 °C fo r 8 h . Th en t h e so lven t s were evap o rat ed an d t h e resid u e ch ro m at o grap h ed o n a co lu m n o f silica gel (150 g, ligh t p et ro leu m / et h yl acet at e 1 : 1 t o 1 : 2) t o give 6 (1.794 g, 76%). M.p . – slo w d eco m p o sit io n o ver 180 °C (ref. 13 gives m .p . 160–162 °C, n o sp ect ral d at a were given ). EI MS, m /z (rel%): 236 (36) [M], 221 (100). 1 H NMR (400 MHz, CDCl 3 ): 0.19 (s, 9 H, SiMe 3 ); 3.15, 3.30 (2 × s, 2 × 3 H, N-Me); 8.16 (s, H-6). 13 C NMR (100 MHz, CDCl 3 ): –0.15 (SiMe 3 ); 27.68, 36.57 (N-Me); 96.03, 96.42, 98.11 (C≡C an d C-5); 148.81 (CH-6); 150.47, 161.20 (C=O -2 an d C=O -4).

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1232 5-Et h yn yl-1,3-d im et h ylp yrim id in e-2,4-(1H,3H)-d io n e (4 )

A 1 M so lu t io n o f TBAF·3H 2 O in THF (10 m l, 10 m m o l) was ad d ed t o silyl d erivat ive 6 (470 m g, 2 m m o l) at 0 °C an d t h e m ixt u re was st irred at ro o m t em p erat u re fo r 5 h . Th en t h e so lven t was evap o rat ed , t h e resid u e was ap p lied o n t o a co lu m n o f silica gel (50 g) an d t h e p ro d u ct was elu t ed wit h a grad ien t o f et h yl acet at e/ MeO H (10 : 0 t o 8 : 2). Th e cru d e p ro d u ct w a s cryst a llized fro m d ich lo ro m et h a n e/ h ep t a n e. Yield 1 6 6 m g (5 1 % ); m .p . 218–221 °C. EI MS, m /z (rel.%): 164 (100) [M], 107 (25), 79 (28), 66 (15), 42 (93). 1 H NMR (400 MHz, CDCl 3 ): 3.17 (s, 1 H, ≡CH); 3.38, 3.42 (2 × s, 2 × 3 H, 2 × CH 3 ); 7.50 (H-6). 13 C NMR (100 MHz, CDCl 3 ): 28.38, 37.37 (N-CH 3 ); 74.92 (C≡); 81.52 (≡CH); ca 142.5 (very weak, C-5); 146.39 (CH-6); 150.85 (C=O -2); 161.74 (C=O -4). Fo r C 8 H 8 N 2 O 2 (164.2) calcu lat ed : 58.53% C, 4.91% H, 17.06% N; fo u n d : 58.79% C, 4.98% H, 16.74% N. Ni-Cat alyzed Cyclo t rim erizat io n s o f Acet ylen es. Gen eral Pro ced u re Method A: A 0.065 M so lu t io n o f Ni(CO D) 2 in THF (6 m l, 0.4 m m o l) was ad d ed d ro p wise t h ro u gh a sep t u m t o a st irred so lu t io n o f an acet ylen e (500 m g, 2 m o l) an d PPh 3 (262 m g, 1 m o l) in TH F (10 m l) u n d er argo n (CAUTIO N! exo t h erm ic react io n ). Th e m ixt u re was st irred at ro o m t em p erat u re fo r 5 h an d t h en t h e so lven t was evap o rat ed . Pro d u ct s were sep arat ed by co lu m n ch ro m at o grap h y o f t h e resid u e o n silica gel (50 g, ligh t p et ro leu m / et h yl acet at e/ m et h an o l (1 : 1 : 0 → 0 : 1 : 0 → 0 : 9 : 1)). Fo r yield s o f t h e p art icu lar p ro d u ct s, see Table I. Meth od B: TH F (1 0 m l) w as ad d ed t o an argo n p u rged flask co n t ain in g acet ylen e 3 a (1 m m o l), NiCp 2 (76 m g, 0.4 m m o l) an d PPh 3 (420 m g, 1.6 m m o l). Th e m ixt u re was st irred at ro o m t em p erat u re o vern igh t . Th e wo rk-u p an d iso lat io n was p erfo rm ed in t h e sam e way as in m et h o d A t o affo rd p ro d u ct 7 a in 35% yield . 1,3,4-T ris(9-benz ylpurin-6-yl)benz ene (7 a). Yello w m icro cryst als; m .p . 155–158 °C (Et O H/ t o lu en e). FAB MS, m /z (rel.%): 703 (15) [M + H], 91 (100). 1 H NMR (500 MHz, CDCl 3 ): 5.40 (s, 4 H, CH 2 Ph ); 5.49 (s, 2 H, CH 2 Ph ); 7.24–7.36 (m , 15 H, H-Ph ); 7.84, 7.88, 8.11 (3 × s, 3 × 1 H, H-8-Pu ); 8.47 (d , 1 H, J = 8.2, H-6-b en zen e); 8.69, 8.73, 9.08 (3 × s, 3 × 1 H, H-2-Pu ); 9.20 (d d , 1 H, J = 8.2, 1.8, H-5-b en zen e); 9.64 (d , 1 H, J = 1.6, H-3-b en zen e). 13 C NMR (100 MHz, CDCl 3 ): 47.15 (CH 2 ); 127.70, 128.47, 129.06, 131.06, 132.52, 133.23 (CH-aro m .); 131.93, 135.19, 136.14, 136.94, 137.79 (CH-aro m . an d C-5); 144.16, 144.50 (CH-8); 151.96, 152.54 (CH-2); 151.71, 153.50, 157.36, 157.94 (C-4 an d C-6). FAB HRMS, fo u n d : 703.2778; C 4 2 H 3 1 N 1 2 [M + H ] req u ires: 7 0 3 .2 7 9 5 . Fo r C 4 2 H 3 0 N 1 2 ·1 / 2 t o lu en e (7 4 8 .8 ) ca lcu la t ed : 72.98% C, 4.58% H, 22.45% N; fo u n d : 72.65% C, 4.55% H, 22.26% N. 1,2,4-T ris[9-(tetrahydropyran-2-yl)purin-6-yl]benzene (7 b). Yello w am o rp h o u s so lid . FAB MS, m /z (rel.%): 685 (9) [M + H], 601 (3) [M + H – THP], 517 (7) [M + 2 H – 2 THP], 433 (100) [M + 3 H – 3 THP], 85 (42) [THP]. 1 H NMR (500 MHz, CDCl 3 , p art icu lar p u rin e rin gs d esign at ed as A, B an d C): 1.60–2.20 (m , 18 H, CH 2 ); 3.80, 4.19 (2 × m , 6 H, CH 2 O ); 5.79 (d , 2 H, J = 9.6, O CHN); 5.88 (d , 1 H, J = 9.9, O CHN); 8.14 (s, 1/ 2 H), 8.16 (s, 1 H) an d 8.19 (s, 1/ 2 H, H-8-Pu A,B); 8.36 (s, 1 H, H-8-Pu C); 8.46 (d , 1 H, J = 8.2, H-6-b en zen e); 8.68, 8.70, 8.72, 8.74 (4 × s, 4 × 1/ 2 H, H-2-Pu A,B); 9.07 (s, 1 H, H-2-Pu C); 9.21 (d , 1 H, J = 8.2, H-5-ben zen e); 9.60 (s, 1 H, H-3-b en zen e). 13 C NMR (100 MHz, APT, CDCl 3 ): 23.51, 25.60, 32.43, 32.58 (4 × CH 2 ); 69.48 (CH 2 O ); 82.60, 82.67, 82.76 (O CHN); 131.87 (C-5-ben zen e); 1 3 2 .2 5 , 1 3 2 .8 1 (C -5 -Pu A,B,C ); 1 3 3 .3 7 (C -6 -b en zen e); 1 3 4 .1 1 (C -3 -b en zen e); 1 3 6 .8 8 (C -2 -b en zen e); 1 3 7 .6 7 (C -4 -b en zen e); 1 3 8 .5 4 (C -1 -b en zen e); 1 4 2 .8 4 , 1 4 2 .9 0 , 1 4 3 .1 2 (C -8 -Pu A,B,C ); 1 5 1 .7 9 (C -4 -Pu A,B,C ); 1 5 2 .5 8 (C -2 -Pu A,B); 1 5 3 .1 3 (C -2 -Pu C ); 1 5 4 .3 6

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(C-6-Pu C); 158.13 (C-6-Pu A); 158.77 (C-6-Pu B). FAB HRMS, fo u n d : 685.3128; C 36 H 37 N 12 O 3 [M + H] req u ires: 685.3112. 1,3,5-T ris[9-(tetrahydropyran-2-yl)purin-6-yl]benzene (8 b). Yello w am o rp h o u s so lid . FAB MS, m /z (rel.%): 685 (27) [M + H], 601 (12) [M + H – THP], 517 (13) [M + 2 H – 2 THP], 433 (100) [M + 3 H – 3 THP]. 1 H NMR (400 MHz, CDCl 3 ): 1.65–2.22 (m , 18 H, CH 2 ); 3.84 (d t , 3 H, J = 2.2, 11.5, CH 2 O a); 4.22 (d , 3 H, J = 11.3, CH 2 O b); 5.90 (d d , 3 H, J = 10.2, 2.4, O CHN); 8.43 (s, 3 H, H-8-Pu ); 9.15 (s, 3 H, H-2-Pu ); 10.36 (s, 3 H, H-ben zen e). 13 C NMR (100 MHz, APT, CDCl 3 ): 23.54, 25.64, 32.65 (3 × CH 2 ); 69.55 (CH 2 O ); 82.77 (O CHN); 132.21 (C -5 -Pu ); 1 3 4 .5 1 (C H -b en zen e); 1 3 7 .5 1 (C -b en zen e); 1 4 3 .2 9 (C -8 -Pu ); 1 5 2 .6 2 (C -6 -Pu ); 153.33 (C-2-Pu ); 155.13 (C-4-Pu ). FAB HRMS, fo u n d : 685.3135; C 36 H 37 N 12 O 3 [M + H] req u ires: 685.3112. 1 ,3 ,4 -T ris(9 -m eth ylpurin -6 -yl)ben z en e (7 c ). Yello w ish p o w d er; m .p . 1 9 5 –1 9 9 °C (Et O H / t o lu en e). EI MS, m /z (rel.%): 474 (18) [M], 446 (17), 210 (15), 129 (35), 57 (100). 1 H NMR (500 MHz, CDCl 3 ): 3.87 (s, 6 H, CH 3 ); 3.95 (s, 3 H, CH 3 ); 7.91 (d , 1 H, J = 9.5, H-b en zen e); 8.12, 8.20, 8.67, 8.72, 9.07, 9.59 (6 × s, 6 × 1 H, H-Pu ); 8.43 (d , 1 H, J = 7.9, H-b en zen e); 9.16–9.21 (m , 1 H, H-ben zen e). 13 C NMR (100 MHz, CDCl 3 ): 29.74, 29.86 (CH 3 ); 131.15, 1 3 2 .5 7 , 1 3 3 .2 3 (C H -b en zen e); 1 3 1 .3 2 , 1 3 1 .8 8 , 1 3 1 .9 3 (C -5 -Pu ); 1 3 6 .1 4 , 1 3 6 .9 6 , 1 3 7 .8 1 (C-b en zen e); 144.97, 145.17, 145.36 (CH-8-Pu ); 151.85, 151.93, 152.45 (CH-2-Pu ); 152.01, 152.09, 153.03, 153.46, 157.30, 157.89 (C-4 an d C-6). EI HRMS, fo u n d : 474.1759; C 24 H 18 N 12 [M] req u ires: 474.1777. 1 ,2 ,4 -T ris(1 ,3 -dim eth yl-2 ,4 -diox o-1 ,2 ,3 ,4 -tetrah ydropyrim idin -5 -yl)ben z en e (9 ). O b t ain ed in 43% yield fro m 4 (react io n t im e 12 h ). Yello w am o rp h o u s so lid . FAB MS, m /z (rel.%): 493 (21) [M + H], 279 (30), 75 (100). 1 H NMR (500 MHz, CDCl 3 ): 3.25 (s, 3 H); 3.29 (s, 3 H); 3.30 (s, 3 H); 3.32 (s, 6 H); 3.40 (s, 3 H, all CH 3 ); 7.30 (d , 1 H, J = 7.8, H-aro m .); 7.51 (s, 1 H, H -6 -U); 7 .6 0 –7 .6 4 (m , 3 H , 2 × H -a ro m . a n d H -6 -U); 8 .0 5 (s, 1 H , H -6 -U). 1 3 C N M R (125.8 MHz, CDCl 3 ): 27.63, 27.72, 36.30, 36.32, 36.47 (all CH 3 ); 110.73, 111.86, 112.21, 132.45, 132.73, 133.39, 150.87, 151.09, 161.54, 161.60, 161.66 (C-aro m .); 127.03, 130.29, 130.66, 142.64, 142.85 (CH-aro m .). FAB HRMS, fo u n d : 493.1864; C 2 4 H 2 5 N 6 O 6 [M+H] req u ires: 493.1836. Co -cyclo t rim erizat io n o f 3 a an d 4 Co -cyclo t rim erizat io n o f 3 a (234 m g, 1 m m o l) an d 4 (164 m g, 1 m m o l) in p resen ce o f Ni(CO D) 2 an d PPh 3 was p erfo rm ed in t h e sam e way as d escribed ab o ve (m et h o d A). A co lu m n ch ro m at o grap h y gave 7 a (59 m g, 25%), u n react ed 4 (49 m g, 30%) an d a co m p lex m ixt u re o f o t h er p ro d u ct s wh ich was re-ch ro m at o grap h ed t o give co m p o u n d 1 0 (16 m g, 5%). 1,2-Bis(9-benz ylpurin-6-yl)-4-(1,3-dim ethyl-2,4-dioxo-1,2,3,4-tetrahydropyrim idin-5-yl)benz ene (1 0 ). Yello wish am o rp h o u s so lid . FAB MS, m /z (rel.%): 633 (9) [M + H], 279 (12), 91 (100). 1 H NMR (500 MHz, CDCl 3 ): 3.43, 3.48 (2 × s, 2 × 3 H, CH 3 -U); 5.39 (s, 4 H, CH 2 Ph ); 7.25–36 (m , 10 H, H-aro m .); 7.54 (s, 1 H, H-6-U); 7.83, 7.85 (2 × s , 2 × 1 H, H-8-Pu ); 7.95 (d d , 1 H, J = 8 .2 , 1 .3 , H -5 -b en zen e); 8 .2 9 (d , 2 H , J = 8 .2 , H -6 -b en zen e); 8 .3 1 (d , 2 H , J = 1 .3 , H-3-b en zen e); 8.65, 8.70 (2 × s, 2 × 1 H, H-2-Pu ). 13 C NMR (125.8 MHz, CDCl 3 ): 28.28, 37.20 (CH 3 ); 47.19 (CH 2 Ph ); 113.23 (C-5-U); 127.73, 128.52, 129.09 (CH-aro m .-Bn ); 129.53 (C -5 -b en zen e); 1 3 1 .3 2 (C -3 -b en zen e); 1 3 1 .7 7 (C -5 -Pu ); 1 3 4 .5 2 , 1 3 5 .0 0 , 1 3 5 .2 1 , 1 3 5 .9 1 (C -a ro m .); 1 4 1 .1 9 (C H -6 -U); 1 4 4 .1 2 (C H -8 -Pu ); 1 5 1 .7 1 (C =O -2 -U a n d C -4 -Pu ); 1 5 2 .0 1 (C H -2 -Pu ); 1 5 7 .4 2 , 1 5 7 .8 5 (C -6 -Pu ); 1 6 1 .9 7 (C =O -4 -U). FAB H RM S, fo u n d : 6 3 3 .2 4 5 7 ; C 36 H 29 N 10 O 2 [M + H] req u ires: 633.2475.

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1,2,4-Tris(p u rin -6-yl)ben zen e (7 e) D o w ex 5 0 X8 (H + fo rm , 5 0 m g) w a s a d d ed t o a so lu t io n o f co m p o u n d 7 b (1 2 0 m g, 0 .1 8 m m o l) in 9 6 % aq u eo u s Et O H (2 0 m l) an d t h e m ixt u re was reflu xed fo r 3 h (TLC sh o wed co m p let io n o f t h e react io n ). Th en t h e resin was filt ered o ff an d wash ed wit h h o t Et O H (20 m l), sat u rat ed et h an o lic am m o n ia (10 m l) an d Et O H (10 m l). Th e co llect ed filt rat es were evap o rat ed an d t h e resid u e cryst allized fro m Et O H. Yield 70 m g (90%). W h it e m icro cryst als; m .p . 319–322 °C. FAB MS, m /z (rel.%): 433 (100) [M + H]. 1 H NMR (500 MHz, DMSO -d 6 ): 8.36 (d , 1 H, J = 8.1, H-6′′); 8.40 (s, 1 H, H-8A); 8.45 (s, 1 H, H-8B); 8.51 (s, 1 H, H-2A); 8.59 (s, 1 H, H-8C); 8.61 (s, 1 H, H-2B); 8.97 (s, 1 H, H-8C); 9.21 (d d , 1 H, J = 1.3, 8.1, H-5′′); 9.50 (d , 1 H, J = 1.2, H-3′′). 13 C NMR (125 MHz, DMSO -d 6 ): 129.27 (C-5A,B); 129.85 (C-5′′); 130.64 (C-5C); 132.00 (C-6′′); 132.19 (C-3′′); 136.29 (C-2′′); 137.09 (C-1′′); 137.51 (C-4′′); 145.46 (C-8A); 145.74 (C-8B); 148.12 (C-8C); 149.80 (C-6C); 150.86 (C-2A); 151.10 (C-2B,C); 154.27 (C-6A); 154.61 (C-4A); 154.85 (C-4B); 155.12 (C-6B); 156.46 (C-4C). Fo r C 21 H 12 N 21 ·2Et O H (524.5) calcu lat ed : 57.24% C, 4.61% H, 32.04% N; fo u n d : 57.37% C, 4.25% H, 31.78% N. FAB HRMS, fo u n d : 433.1337; C 21 H 13 N 12 [M + H] req u ires: 433.1386. T his work is a part of a research project Z 4 055 905. It was supported by the Grant Agency of the Czech Republic (grants N o. 203/00/0036 to M. H. and N o. 203/02/0248 to I. S.). T he cytostatic activity was studied by Dr I. V otruba whose contribution is gratefully acknowledged. T he authors’ thanks are also due to Ms K. Havlíčková for excellent technical assistance.

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